Immersion Oil

[Knecht, David]

This has been a fascinating and informative discussion.  Let me add another twist.  If you are imaging at both room temperature and 37C do you use different oils?  I presume that the Zeiss/Nikon oils are formulated for 22C, while Cargille makes one specifically for 37C applications.  Dave

On Mar 17, 2009, at 4:49 PM, S. Brunet wrote:

Hello:

Don't mix Zeiss Immersol 518F and Cargille Type 37 immersion oils (the SOP I
wrote was clear on this!).  I tried cleaning it but I had no luck.  If I put a
drop of Zeiss oil on that objective and put a coverslip on, I can clearly see a
region which does not mix when I image the oil area/volume under a different
microscope!

I clean all objectives between batches of Zeiss oil just in case.

Bye!
Sophie
____________________________________________________
Sophie M. K. Brunet, Ph. D.
Research Officer
Optical Spectroscopy, Laser Systems and Applications
[hidden email]
306-966-1719 (office)   306-966-1702 (fax)
____________________________________________________
Saskatchewan Structural Sciences Centre
University of Saskatchewan
Thorvaldson Bldg.
110 Science Place
Saskatoon, Sk   S7N 5C9
____________________________________________________


Quoting David Knecht <[hidden email]>:

We now have Nikon, Zeiss and Leica microscopes in our facility.  Is

the problem of cross-contamination of immersion oil real or

hypothetical?   Is it possible to use a single type of immersion oil

for all, so that we don't have to worry about moving samples between

microscopes?   Dave

Dr. David Knecht

Department of Molecular and Cell Biology

Co-head Flow Cytometry and Confocal Microscopy Facility

U-3125

91 N. Eagleville Rd.

University of Connecticut

Storrs, CT 06269

860-486-2200

860-486-4331 (fax)

Dr. David Knecht    

Department of Molecular and Cell Biology

Co-head Flow Cytometry and Confocal Microscopy Facility

U-3125

91 N. Eagleville Rd.

University of Connecticut

Storrs, CT 06269

860-486-2200

860-486-4331 (fax)

[Dale Callaham]

Hi All,

The oils definitely have different compositions and viscosities and give
strange optical results when people do manage to mix them. We have used
Cargille oils (Type DF primarily) for general fluorescene work for the
15 yr I've been in current position and no lens has been damaged (by the
oil); we just wipe excess from oil lenses and do not clean with solvent
ever (unless they got the mixture treatment or other contamination) and
have not had any lens issues. We mostly have Nikon scopes aside from the
Zeiss 510 confocal. As a multiuser facility we can't control everything
so I routinely find little bottles of Nikon oil near the scopes and
dispose of them directly; nothing against Nikon oil - we just try to
avoid mixing - the point of this being that the objectives get dunked in
just about everything and seem to survive.  We had one oil objective
fail on a Zeiss inverted system - front element just dropped into the
lens barrel - but suspect it was just a chance event - nothing like it
has happened since. On that scope we use the Zeiss Immersol 518F - Zeiss
has a data sheet on their website and has issued a bulletin on it; when
cool it formed crystals so we keep in in a 35C incubator always. They
may have fixed the issue, but it is definitely not a single component oil.

Cheers

S. Brunet wrote:

> Hello:
>
> Don't mix Zeiss Immersol 518F and Cargille Type 37 immersion oils (the SOP I
> wrote was clear on this!).  I tried cleaning it but I had no luck.  If I put a
> drop of Zeiss oil on that objective and put a coverslip on, I can clearly see a
> region which does not mix when I image the oil area/volume under a different
> microscope!
>
> I clean all objectives between batches of Zeiss oil just in case.
>
> Bye!
> Sophie
> ____________________________________________________
> Sophie M. K. Brunet, Ph. D.
> Research Officer
> Optical Spectroscopy, Laser Systems and Applications
> [hidden email]
> 306-966-1719 (office)   306-966-1702 (fax)
> ____________________________________________________
> Saskatchewan Structural Sciences Centre
> University of Saskatchewan
> Thorvaldson Bldg.
> 110 Science Place
> Saskatoon, Sk   S7N 5C9
> ____________________________________________________
>
>
> Quoting David Knecht <[hidden email]>:
>
>> We now have Nikon, Zeiss and Leica microscopes in our facility.  Is
>> the problem of cross-contamination of immersion oil real or
>> hypothetical?   Is it possible to use a single type of immersion oil
>> for all, so that we don't have to worry about moving samples between
>> microscopes?   Dave
>>
>> Dr. David Knecht
>> Department of Molecular and Cell Biology
>> Co-head Flow Cytometry and Confocal Microscopy Facility
>> U-3125
>> 91 N. Eagleville Rd.
>> University of Connecticut
>> Storrs, CT 06269
>> 860-486-2200
>> 860-486-4331 (fax)
>>
>>
>>

[Keith Morris]

Cargille make the very low fluorescence DF immersion oil and the FF
immersion oil. Over 5 years or so ago the DF oil [blue label] used to
clearly state on the label that the oil could damage the objectives when
used very frequently [I'm paraphrasing, that old bottle's back at UCL, and
I'm pretty sure it was the DF not the FF that said it].  

Their FF oil was stated as being perfectly safe in this respect [but it was
hinted it wasn't as superb as DF in terms of auto-fluorescence]. For this
reason I always 'bottled out' from using the DF and stuck with the FF. My
new bottles of DF [and the Cargille website] don't state this anymore so
presumably Cargille have relatively recently modified the oils chemistry and
additives to make it even kinder to objectives. Besides Nikon recommend
Cargille DF, so I use it.

Ironically we used Cargille FF on Zeiss Axiovert 100 microscopes because
back then Zeiss 518F immersion oil had a real problem with going cloudy when
the room temperature went down to 16 oC [in fact it used to set solid at
this temperature and required 'two hours at 40oC in a water bath' (or 10
seconds or so in a microwave) to 're-dissolve' the crystals and clear the
oil.

Zeiss has since modified 518F so that this is far less of a problem, but
crystals can still form below 18oC, particularly if say the sample
slide+518F oil on the cover-slip is stored in a fridge. However Cargille oil
FF and DF in comparison has a cloud point of -6oC [far far lower than
Zeiss's 518F oil] - hence the reason we had to use Cargille FF in the cold
lab [room air conditioner problems as usual].

Even now Zeiss state for their D518F oil: "At temperatures below 18°C (65°F)
crystal formation may occur in Immersol 518F & 518N immersion oils (both
during shipping or regular storage). This is due to the isomerism of the
main component Di-(TCD?methylol adipate (DTCDMA) and our demands on the
purity.".. They go on to discuss how to dissolve the crystals [see above].

This very different cloud point temperature alone suggests that the two
manufacturer's oils have very different chemical characteristics and won't
mix well. They have different things dissolved in them, and neither are a
pure 'oil' as such. I expect mixing will simply precipitate those crystals
in the Zeiss 518F oil. That said all Cargille immersion oils are designed to
be miscible with each other. Likewise oil acidity [it's not got a pH] will
increase with storage [safe storage times typically range from 5 to 10
years, and initial acidity is very low]. Immersion oils are also matched and
tested with a manufacturers lens 'cement' to minimise damage to the
objectives as much as for their optical imaging properties, so it probably
pays to use the manufacturer's recommended immersion oil [Cargille DF with
Nikon and Zeiss 518F with Zeiss, although I believe the Leica immersion oil
is identical to the Zeiss [might be wrong].

I attach a few threads from the microscopy.com list-server immersion oil
postings [2004] below that seem relevant.

Immersion oils don't 'dry out' or harden at all in normal use [Cargille
developed the first non-drying immersion oil back in the 1940s]. So if the
microscope is in frequent use, removing any oil residue with solvents is
unnecessary [unless you've mixed non-miscible types or more likely the
immersion oil is contaminated with say Vectashield, nail varnish, culture
media or pen marker ink. As solvents used to remove immersion oil are more
implicated with damage to the lens cement than the oils [particularly
ethanol, I only use Ether], it's probably best to stick to the recommended
oil for the microscope and just gently wipe the excess away between use with
a dry lens tissue - simply to prevent the immersion oil from dripping into
the works.

I've never had any problems using Zeiss or Cargille immersion oils at 37oC
with live cell work, although the oils are optimised for room temperature
use.

Keith

http://www.2spi.com/catalog/ltmic/cargille.shtml

http://www.microscopy-uk.org.uk/mag/artfeb04/cdclean.html

http://www.ibms.org/pdf/lens_cleaning.pdf


---------------------------------------------------------------------------
Dr Keith J. Morris,
Molecular Cytogenetics and Microscopy Core, Laboratory 00/069 and 00/070,
The Wellcome Trust Centre for Human Genetics, Roosevelt Drive, Oxford  OX3
7BN, United Kingdom.

Telephone:  [phone]
Email:  [hidden email]
Web-pages: http://www.well.ox.ac.uk/cytogenetics/









----------------------------------------------------------------------------
Greetings,
In connection to the thread, at one time in the past
different immersion oils from the different 'scope makers had
diferent refractive indices. Therefore, if they got mixed, the
results were really bad, and presumably the lens would have been
designed around a given index. I *think* that the oils from Zeiss,
Nikon, Olympus, Leica now have the same refractive index, but I am
not sure, and I would be interested to hear if anyone knows.

Tobias Baskin
}
}
} The last part of the letter listed below indicates damage to Nikon
} lenses by the use of Zeiss immersion oil. Do you, Ralph, or anyone
} else have instances where this has occurred? (I would suspect that
} this would occur on inverted scopes most often and would be the
} result of dissolving the seal between the lens and the outer case.)
} If so was this the old Zeiss or the new Zeiss oil immersion medium?
} I've got Nikon scopes and would like to post messages in the
} appropriate rooms if this has proven to be a problem. I also have
} Zeiss oil over 15 years old with no solidification problems that
} I've kept hidden.
}
} Also, does anyone know where to find small plastic 5-10 ml bottles
} that would be suitable for dispensing oil immersion medium?
}
} .. Jerry Calvin
}
} listed 3-3-04
} .. Also be aware that different types of immersion oil should never
} be allowed to mix, and that some types of oil can damage the
} mounting cement of some brands of lenses. Using Zeiss immersion oil
} with some Nikon lenses, for example, can be disastrous.

--
_ ____ __ ____
/ \ / / \ / \ \ Tobias I. Baskin
/ / / / \ \ \ Biology Department
/_ / __ /__ \ \ \__ 611 N. Pleasant St.
/ / / \ \ \
University of Massachusetts
/ / / \ \ \
Amherst, MA, 01003
/ / ___ / \ \__/ \ ____ Voice: 413 - 545 - 1533
Fax: 413 - 545 - 3243
http://www.bio.umass.edu/biology/baskin/







From: Jim Sanzo :      jsanzo-at-acrionline.org


----------------------------------------------------------------------------
----

Date: Wed, 17 Mar 2004 15:52:25 -0500
Subject: [Microscopy] immersion oils


----------------------------------------------------------------------------
----

Contents Retrieved from Microscopy Listserver Archives
http://www.microscopy.com/MicroscopyListserver/MicroscopyArchives.html


----------------------------------------------------------------------------
----



There is yet another issue: miscibility.

Some "oils" are not oils at all, at least not in the sense that they are
derived from petroleum, or contain fully saturated long-carbon chain
(aliphatics/alkanes, etc) compounds. Virtually any substance that has the
correct RI, with useable physical properties will be suitable as an
immersion "oil". Unfortunately, this leaves the door wide open for
manufacturers to use anything from a "real" oil, to compounds like benzyl
benzoate, or mixtures of a liquid with a dissolved solid component. The
latter case is probably what has caused the infamous Zeiss crystallizing-oil
problem. The Zeiss oil is probably saturated when it comes from the factory.
When it experiences a drop in temperature, or a bit of evaporation, the
solution becomes supersaturated and the crystals drop out. As has been
mentioned, warming the mix redissolves the crystals. At that point, I would
imagine the RI of the oil would be too high, and would create spherical
aberration.

In any case, if you mix oils on the same microscope slide or objective lens
without thorough cleaning, chances are you'll create a blurry liquid-liquid
interface, and there goes the resolution! Since all the microscope
manufacturers know that their oil might not be miscible with any other
manufacturers oil, they always recommend that you pick one oil and use it
exclusively.

Jim


-----Original Message-----
} From: Tobias Baskin [mailto:baskin-at-bio.umass.edu]
Sent: Wednesday, March 17, 2004 1:50 PM
To: microscopy-at-MSA.microscopy.com

Greetings,
In connection to the thread, at one time in the past
different immersion oils from the different 'scope makers had
diferent refractive indices. Therefore, if they got mixed, the
results were really bad, and presumably the lens would have been
designed around a given index. I *think* that the oils from Zeiss,
Nikon, Olympus, Leica now have the same refractive index, but I am
not sure, and I would be interested to hear if anyone knows.

Tobias Baskin
}
}
} The last part of the letter listed below indicates damage to Nikon
} lenses by the use of Zeiss immersion oil. Do you, Ralph, or anyone
} else have instances where this has occurred? (I would suspect that
} this would occur on inverted scopes most often and would be the
} result of dissolving the seal between the lens and the outer case.)
} If so was this the old Zeiss or the new Zeiss oil immersion medium?
} I've got Nikon scopes and would like to post messages in the
} appropriate rooms if this has proven to be a problem. I also have
} Zeiss oil over 15 years old with no solidification problems that
} I've kept hidden.
}
} Also, does anyone know where to find small plastic 5-10 ml bottles
} that would be suitable for dispensing oil immersion medium?
}
} .. Jerry Calvin
}
} listed 3-3-04
} .. Also be aware that different types of immersion oil should never
} be allowed to mix, and that some types of oil can damage the
} mounting cement of some brands of lenses. Using Zeiss immersion oil
} with some Nikon lenses, for example, can be disastrous.

--
_ ____ __ ____
/ \ / / \ / \ \ Tobias I. Baskin
/ / / / \ \ \ Biology Department
/_ / __ /__ \ \ \__ 611 N. Pleasant St.
/ / / \ \ \
University of Massachusetts
/ / / \ \ \
Amherst, MA, 01003
/ / ___ / \ \__/ \ ____ Voice: 413 - 545 - 1533
Fax: 413 - 545 - 3243
http://www.bio.umass.edu/biology/baskin/

 

[Martin Spitaler]

I was told by a Leica engineer a few years ago that Leica and Zeiss buy
their immersion oil from the same supplier, and indeed the specifications in
the data sheet are identical. We have ever since used the same oil from
Leica or Zeiss (whatever is cheaper) on all microscopes without prolems,

Martin

######################################
Martin Spitaler, PhD

FILM - Facility for Imaging by Light Microscopy
- Facility Manager -
Sir Alexander Fleming Building, desk 401
Imperial College London / South Kensington
Exhibition Road
London SW7 2AZ
UK

Tel. [phone]
E-mail [hidden email]
Website: http://imperial.ac.uk/imagingfacility

[Monique Vasseur]

I had been said that Zeiss immersion oils contain a small amount of solvent in it in order to dissolve old dried oil that could have stayed on the objective. Can someone comment on this if this is true?

Monique Vasseur
tél. [phone] poste 5148
-----Message d'origine-----
De : Confocal Microscopy List [mailto:[hidden email]] De la part de S. Brunet
Envoyé : 17 mars 2009 16:50
À : [hidden email]
Objet : Re: Immersion Oil

Hello:

Don't mix Zeiss Immersol 518F and Cargille Type 37 immersion oils (the SOP I
wrote was clear on this!).  I tried cleaning it but I had no luck.  If I put a
drop of Zeiss oil on that objective and put a coverslip on, I can clearly see a
region which does not mix when I image the oil area/volume under a different
microscope!

I clean all objectives between batches of Zeiss oil just in case.

Bye!
Sophie
____________________________________________________
Sophie M. K. Brunet, Ph. D.
Research Officer
Optical Spectroscopy, Laser Systems and Applications
[hidden email]
306-966-1719 (office)   306-966-1702 (fax)
____________________________________________________
Saskatchewan Structural Sciences Centre
University of Saskatchewan
Thorvaldson Bldg.
110 Science Place
Saskatoon, Sk   S7N 5C9
____________________________________________________


Quoting David Knecht <[hidden email]>:

> We now have Nikon, Zeiss and Leica microscopes in our facility.  Is
> the problem of cross-contamination of immersion oil real or
> hypothetical?   Is it possible to use a single type of immersion oil
> for all, so that we don't have to worry about moving samples between
> microscopes?   Dave
>
> Dr. David Knecht
> Department of Molecular and Cell Biology
> Co-head Flow Cytometry and Confocal Microscopy Facility
> U-3125
> 91 N. Eagleville Rd.
> University of Connecticut
> Storrs, CT 06269
> 860-486-2200
> 860-486-4331 (fax)
>
>
>

[Edelmann, Richard E. Dr.]

I am going to chime in agreement with bad results from mixing oils.  
About six years ago we ran into problems with mixing oils (Fryer oil
which was a repackaged Cargille vs Olympus, etc.) and had severe
issues with resolution degradation.  And we found that it took
serious scrupulous cleaning the lenses to remove all contamination.  
(No gooy oil or concretions, etc. just hard to scrupulously clean).

Yes, scrupulously cleaning previous oil from slides "could have
worked" but the fact is users rearely clean the old oils well enough.

 Alternately  requiring slides not to be used with oil anywhere else -

 but that also is a trust system. So rather than waste time cleaning
lenses, I simply bought a bunch of new oil and went around and traded

oil bottle with every lab.  Since I was giving them new oil no one
complained.  No more mixing issues.

A little expensive maybe but time and good faith it was worth it.


On 17 Mar 2009 at 13:43, Carl Boswell wrote:

> The stories are real, at least with respect to the miscibility of
> different oils.  While one might think that oil is oil, two different
> immersion oils together can behave like oil and water.  That is why I
> insist that samples first viewed on some other microscope be
> scrupulously cleaned of whatever oil was used before applying oil used
> with our Core systems.
>
> As for the issue of one manufacture's oil eating away another
> manufacture's lens, I'd consider the source of that information
> carefully.  I also use Cargille oil on multiple sytems and have no
> complaints. C
>
> Carl A. Boswell, Ph.D.
> Molecular and Cellular Biology
> University of Arizona
> 520-954-7053
> FAX [phone]
> ----- Original Message -----
> From: David Knecht<mailto:[hidden email]>
> To: [hidden email]<mailto:[hidden email]>
> Sent: Tuesday, March 17, 2009 6:56 AM
> Subject: Immersion Oil
>
> We now have Nikon, Zeiss and Leica microscopes in our facility.  Is the problem of cross-contamination of immersion oil real or hypothetical?   Is it possible to use a single type of immersion oil for all, so that we don't have to worry about moving samples between microscopes?   Dave
>
> Dr. David Knecht
> Department of Molecular and Cell Biology
> Co-head Flow Cytometry and Confocal Microscopy Facility
> U-3125
> 91 N. Eagleville Rd.
> University of Connecticut
> Storrs, CT 06269
> 860-486-2200
> 860-486-4331 (fax)
>
>

Richard E. Edelmann, Ph.D.
EXPO Editor, Microscopy and Microanalysis Supplement
Electron Microscopy Facility Director
364 Pearson Hall
Miami University, Oxford, OH 45056
Ph: 513.529.5712        Fax: [phone]
E-mail: [hidden email]
http://www.emf.muohio.edu

[Jurriaan Zwier]

Hello,

I have measured a couple of years back the detoriation of the point spread
function of a confocal microscope upon changing the f of the oil by small
steps of 0.002. The PSF deteriorates very rapidly. I can confirm mixing of
oils is not a good idea. Using oil developed for measurements at 23
degrees for live cell measurements at 37 degrees is a bad idea as well.
For those of you interested, some observations regarding the psf's using
different immersion oils was recently published:

'Immersion oil for high-resolution live-cell imaging at 37ħC: optical and
physical characteristics', L.C.J.M. Oomen, R. Sacher, H.H.J. Brocks, J. M.
Zwier, G. J. Brakenhoff and K. Jalink, Journal of Microscopy 232 (2008)
p353-361

Kind regards,
Jurriaan
-no commercial interest

> When you are down to the third decimal place it's not going to make a huge
> difference*.  You might want to try and see if you observe any difference
> between oil, but I suspect you won't see much change.
>
> Craig
>
> *unless you are talking about fiber optics or some optoelectronics...
>
>
> On Tue, Mar 17, 2009 at 9:58 AM, stu_the_flat
> <[hidden email]>wrote:
>
>> Isn't there a 0.002 difference in the diffraction between some of the
>> immersion oils?
>>
>> Off the top of my head
>>
>> Olympus Oil = 1.515
>> Ziess Oil = 1.515
>> Nikon Oil = 1.513
>>
>> I don¡¦t think it make the world of difference but as I am interested in
>> PSFs
>> I always match the oil the microscope.
>>
>> Stuart
>>
>>
>>
>>
>> David Knecht-charter wrote:
>> >
>> > Being a natural skeptic, that has been my presumption as well.  I
>> > haven't talked to anyone who has actually seen mixed oils gum up their
>> > scope or dissolve  lens cement (if you believe that last one, I have a
>> > few million dollars in my uncle's  account in Africa I will be happy
>> > to send you).  Dave
>> >
>> > On Mar 17, 2009, at 11:37 AM, Peter Carroll wrote:
>> >
>> >>
>> >> > I have been told by a Nikon representative that Zeiss oil could
>> >> destroy the Nikon objective lens
>> >>
>> >> Of course you heard that... Nikon wants you to purchase /their/
>> >> consumables, not their competitors! Heh...
>> >
>> > Dr. David Knecht
>> > Department of Molecular and Cell Biology
>> > Co-head Flow Cytometry and Confocal Microscopy Facility
>> > U-3125
>> > 91 N. Eagleville Rd.
>> > University of Connecticut
>> > Storrs, CT 06269
>> > 860-486-2200
>> > 860-486-4331 (fax)
>> >
>> >
>> >
>> >
>>
>> --
>> View this message in context:
>> http://n2.nabble.com/Immersion-Oil-tp2491973p2492239.html
>> Sent from the Confocal Microscopy List mailing list archive at
>> Nabble.com.
>>
>

[Higdon, Michael]

You definitely have to use different oils.  The refractive index changes.

 

---

From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of David Knecht ATT
Sent: Tuesday, March 17, 2009 9:18 PM
To: [hidden email]
Subject: Re: Immersion Oil

 

This has been a fascinating and informative discussion.  Let me add another twist.  If you are imaging at both room temperature and 37C do you use different oils?  I presume that the Zeiss/Nikon oils are formulated for 22C, while Cargille makes one specifically for 37C applications.  Dave

 

On Mar 17, 2009, at 4:49 PM, S. Brunet wrote:

Hello:

Don't mix Zeiss Immersol 518F and Cargille Type 37 immersion oils (the SOP I
wrote was clear on this!).  I tried cleaning it but I had no luck.  If I put a
drop of Zeiss oil on that objective and put a coverslip on, I can clearly see a
region which does not mix when I image the oil area/volume under a different
microscope!

I clean all objectives between batches of Zeiss oil just in case.

Bye!
Sophie
____________________________________________________
Sophie M. K. Brunet, Ph. D.
Research Officer
Optical Spectroscopy, Laser Systems and Applications
[hidden email]
306-966-1719 (office)   306-966-1702 (fax)
____________________________________________________
Saskatchewan Structural Sciences Centre
University of Saskatchewan
Thorvaldson Bldg.
110 Science Place
Saskatoon, Sk   S7N 5C9
____________________________________________________


Quoting David Knecht <[hidden email]>:

We now have Nikon, Zeiss and Leica microscopes in our facility.  Is

the problem of cross-contamination of immersion oil real or

hypothetical?   Is it possible to use a single type of immersion oil

for all, so that we don't have to worry about moving samples between

microscopes?   Dave

 

Dr. David Knecht

Department of Molecular and Cell Biology

Co-head Flow Cytometry and Confocal Microscopy Facility

U-3125

91 N. Eagleville Rd.

University of Connecticut

Storrs, CT 06269

860-486-2200

860-486-4331 (fax)

 

 

 

 

Dr. David Knecht    

Department of Molecular and Cell Biology

Co-head Flow Cytometry and Confocal Microscopy Facility

U-3125

91 N. Eagleville Rd.

University of Connecticut

Storrs, CT 06269

860-486-2200

860-486-4331 (fax)

 

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[Steffen Dietzel]

At 13:07 18.03.2009, Martin Spitaler wrote:
>I was told by a Leica engineer a few years ago that Leica and Zeiss buy
>their immersion oil from the same supplier,


That's one way to put it: The Leica safety data
sheet for "Immersion oil, without
autofluorescence" (10 ml, 11513859) states as producer: Carl Zeiss, Oberkochen.

Still, Zeiss produces different Oils, (518F,
518N) and this discussion suggested that formulas
may change from time to time. Just to be on the
safe side, I'd rather not mix different batches.
But I'd agree that it is probably worse to have
left over oil on a slide that comes out of the
fridge and condensing water mixing with that.

The technical information also says:
temperature coefficient delta n (theta), range 10/40°C: -0,00038/°C.
I am not 100% sure, but I guess that's the
difference in Ri per degree. So, for a shift from
room temperature (22°C) to 37° that would be a
difference of -0.0057, i.e at 546,1 nm from 1.5180 to 1.5123.

That'd be in the same range you have differences anyway if you use multi-color:
According to the data sheet Ri at 435 nm is
1.5290 and at 643,8 nm it is 1.5124, so between
the two colors the difference is 0.0166 and thus
appears to be much larger than the reported 0.002
between brands (where, presumably both values
were measured at 546 nm - hopefully).

Did I miss anything?

Steffen

>and indeed the specifications in
>the data sheet are identical. We have ever since used the same oil from
>Leica or Zeiss (whatever is cheaper) on all microscopes without prolems,
>
>Martin
>
>######################################
>Martin Spitaler, PhD
>
>FILM - Facility for Imaging by Light Microscopy
>- Facility Manager -
>Sir Alexander Fleming Building, desk 401
>Imperial College London / South Kensington
>Exhibition Road
>London SW7 2AZ
>UK
>
>Tel. [phone]
>E-mail [hidden email]
>Website: http://imperial.ac.uk/imagingfacility

--
---------------------------------------------------------------------------------------------------
Steffen Dietzel, PD Dr. rer. nat
Ludwig-Maximilians-Universität München
Walter-Brendel-Zentrum für experimentelle Medizin (WBex)
Head of light microscopy

Mail room (for letters etc.):
Marchioninistr. 15, D-81377 München

Building location and address for courier, parcel services etc:
Marchioninistr. 27, D-81377 München (Großhadern)

Phone: [phone]
Fax-to-email:   +49/89/2180-9976509
skype: steffendietzel
e-mail: [hidden email]

[Keith Morris]

Hi all,

 

Using immersion oil at 37oC.

 

Zeiss sell a range of low fluorescence immersion oils: bottles of 20, 250 and 500 ml, unfortunately all filled with the same 518F oil. According to Zeiss it’s apparently ‘compatible’ with our Zeiss microscopes, but little extra information is offered on the Zeiss website.

 

The only info I can gleam from the web [other than the ubiquitous 518F safety data Sheet] is the following

Immersion oil "Immersol" 518 F fluorescence free, (ISO 8036-1/2, n(e)=1.518 (23°C), halogen-free)

 

So not much comment about 37oC use.  PeCon’s site [who supply most of Zeiss’s decent incubator stuff] have no opinion on the matter either.

 

The 518F oil safety sheet mentions “Use of the Substance / Preparation: For application information, consult the processing instructions. Processing instructions or technical

information sheet available on request.”. I’ll ask our knowledgeable local Zeiss Confocal rep for more details, as we are just installing a Zeiss/PeCon 37oC live cell incubator here on our Zeiss 510 MetaHead confocal.

Cargilee’s site in comparison is passionate about all things immersion oil:
http://www.cargille.com/immeroil.shtml
http://www.cargille.com/immeroilselection.shtml
and their excellent, if elderly, ‘primer’
http://www.cargille.com/immersionoilmicroscope.shtml

From postings to this server back in 2006, and Cargille’s site not mentioning ‘non-fluorescence’ for type 37, it appears that Cargille 37 probably autofluoresces, so it’s likely better for live cell phase-contrast [halogen lamp] transmission microscopy [but then often so are dry objectives]. I’ll stick with standard 518F immersion oil at 37oC though, it’s always imaged well enough on Zeiss microscopes at that temperature [and at least at 37oC it never became cloudy due to crystal precipitation]. Similarly the Cargille DF and FF seem fine to me at 37oC, and we stick ruthlessly with the same brand on each microscope. A lot of our live cell work is/was done with low power [20x Phase] air objectives though.

I do notice that immersion oil often ingresses into oil immersion objective internal optics with time, although strangely it rarely notices when imaging [like dirt on much of the internal optics, you must mostly focus through it]. One microscopy core manager even commented to me that he considers oil objectives ‘consumables’. The cost of ‘repair’ [often the same price as a new objective] is so high we frequently have to live with it [I advise users that our say 40x objective image quality is suspect for this reason]. In truth we don’t notice any drop in image quality when it must have happened, although you can see it ‘smeared’ on the optics when looking through the removed objective, using a magnifying glass, bright room light and a ‘cleaned’ objective top lens.

Old listerver posting on Cargille 37 attached below.

Regards

Keith

PS missed this link on the last post:

Dissolving crystals in Zeiss 518F oil
http://www.zeiss.com/C1256F8500454979/0/E8CD09DFA520787FC1256F86004B5FFB/$file/immersol-crystals.pdf

---------------------------------------------------------------------------
Dr Keith J. Morris,
Molecular Cytogenetics and Microscopy Core,
Laboratory 00/069 and 00/070,
The Wellcome Trust Centre for Human Genetics,
Roosevelt Drive,
Oxford  OX3 7BN,
United Kingdom.

Telephone:  [phone]
Email:  [hidden email]
Web-pages: http://www.well.ox.ac.uk/cytogenetics/

 

Subject:	Re: Fluorescence from Type 37 Immersion Oil
From:	Vitaly Boyko <[hidden email]>
Reply-To:	Vitaly Boyko <[hidden email]>
Date:	Wed, 16 Aug 2006 10:23:20 -0400
Content-Type:	text/plain

 

Search the CONFOCAL archive at

http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

PS What would be more appropriate to compare is Zeiss 518F (not 518C!) with 

the Nikon type A or Cargille type DF.

Hi Jason,

I wonder why you have tested the Cargille FF imm. oil, as the refractive

index of FF oil is too far off for the standard glass (1.518) ?????!!!!!

Actually, I haven't noticed the "very bad" autofluorescence of the Cargille

type DF versus Zeiss 518 in the CFP, YFP or FRET channels. The difference is

less than 10%.

The problem with the Nikon oil is its dispenser and/or viscosity - it is

often annoying "fighting" with the air bubbles.

What is about the Invitrogen/Mol.Probes mounting media autofluorescence?

I am fed up with the irreproducibility of the background autofluorescence

using the ProlongGold mounting media!!! That is a serious problem compared

to the issue of the immersion oils.

Cheers,

Vitaly

NCI-Frederick

----- Original Message ----- 

From: "Kilgore, Jason" <[hidden email]>

To: <[hidden email]>

Sent: Tuesday, August 15, 2006 7:26 PM

Subject: Re: Fluorescence from Type 37 Immersion Oil

> Search the CONFOCAL archive at

> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

> 

> Back in 2000 I performed a comparison of four different immersion oils

> (Zeiss 518C, Nikon Type A, Cargille Type DF, and Cargille Type FF) for

> autofluorescence (using DAPI, FITC, acridine orange, and TRITC filter

> sets) and general resolution at 60x on a Nikon E400.  I didn't test

> Cargille 37.

> 

> Green autofluorescence tended to be worst for all four, but was least

> for Cargille FF, followed by Nikon Type A, then Zeiss, then (very bad)

> Cargille DF.  However, the resolution was unacceptable for Cargille FF.

> Autofluorescence in the other channels was moderate (DAPI, TRITC) to

> very low (acridine orange), with the same comparative trend applying.

> 

> Thus, ever since we have been using Nikon Type A with no complaints for

> most of our imaging.

> 

> Feel free to contact me personally and I can send you the data.

> 

> Jason

> 

> Jason A. Kilgore

> Cell Biology / Histology

> Molecular Probes/Invitrogen

> [hidden email]

> 

> 

> -----Original Message-----

> From: Confocal Microscopy List [mailto:[hidden email]] On

> Behalf Of S. Brunet

> Sent: Tuesday, August 15, 2006 11:19 AM

> To: [hidden email]

> Subject: Fluorescence from Type 37 Immersion Oil

> 

> Search the CONFOCAL archive at

> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

> 

> Hello to all,

> 

> We recently began using the Cargille 37 oil with the temperature chamber

> on the

> microscope.  I found significant fluorescence from the oil when exposed

> to the

> HBO lamp (I could not see the eYFP because of the blue background).

> 

> So I used the 20x air objective and I was able to obtain an image from

> the

> fluorescence from a drop of 37 oil placed on a coverslip as a sample

> (excitation at 488 or 458nm and pinhole at the usual confocal setting).

> 

> Is this the norm?

> 

> Regards,

> Sophie

> _______________________________________

> Sophie M.K. Brunet, Ph.D.

> Research Officer

> Optical Spectroscopist, Laser Systems and Applications

> [hidden email]

> 306-966-1719 (office)   306-966-1702 (fax)

> _______________________________________

> Saskatchewan Structural Sciences Centre

> University of Saskatchewan

> Thorvaldson Bldg.

> 110 Science Place

> Saskatoon, SK   S7N 5C9

> 

 

[lechristophe]

I would like to point out that Cargille has a new oil for fluorescence
imaging at 37°C, called 37DF, that has very low auto fluorescence. It
is not on their website, but I could get the product pdf sent to me by
a Cargille person, so I can send it if someone is interested. I'm
trying to get some here in France, but none of the french Cargille
distributors has heard about it yet... It has been found to be quite
good for fluorescence imaging of live cells at 37°C, according to this
paper in J Microscopy :

Immersion oil for high-resolution live-cell imaging at 37 degrees C:
optical and physical characteristics.
Oomen LC, Sacher R, Brocks HH, Zwier JM, Brakenhoff GJ, Jalink K.
J Microsc. 2008 Nov;232(2):353-61.


On Mon, Mar 23, 2009 at 11:18, Keith Morris <[hidden email]> wrote:

>
> Hi all,
>
>
>
> Using immersion oil at 37oC.
>
>
>
> Zeiss sell a range of low fluorescence immersion oils: bottles of 20, 250 and 500 ml, unfortunately all filled with the same 518F oil. According to Zeiss it’s apparently ‘compatible’ with our Zeiss microscopes, but little extra information is offered on the Zeiss website.
>
>
>
> The only info I can gleam from the web [other than the ubiquitous 518F safety data Sheet] is the following
>
> Immersion oil "Immersol" 518 F fluorescence free, (ISO 8036-1/2, n(e)=1.518 (23°C), halogen-free)
>
>
>
> So not much comment about 37oC use.  PeCon’s site [who supply most of Zeiss’s decent incubator stuff] have no opinion on the matter either.
>
>
>
> The 518F oil safety sheet mentions “Use of the Substance / Preparation: For application information, consult the processing instructions. Processing instructions or technical
>
> information sheet available on request.”. I’ll ask our knowledgeable local Zeiss Confocal rep for more details, as we are just installing a Zeiss/PeCon 37oC live cell incubator here on our Zeiss 510 MetaHead confocal.
>
> Cargilee’s site in comparison is passionate about all things immersion oil:
> http://www.cargille.com/immeroil.shtml
> http://www.cargille.com/immeroilselection.shtml
> and their excellent, if elderly, ‘primer’
> http://www.cargille.com/immersionoilmicroscope.shtml
>
> From postings to this server back in 2006, and Cargille’s site not mentioning ‘non-fluorescence’ for type 37, it appears that Cargille 37 probably autofluoresces, so it’s likely better for live cell phase-contrast [halogen lamp] transmission microscopy [but then often so are dry objectives]. I’ll stick with standard 518F immersion oil at 37oC though, it’s always imaged well enough on Zeiss microscopes at that temperature [and at least at 37oC it never became cloudy due to crystal precipitation]. Similarly the Cargille DF and FF seem fine to me at 37oC, and we stick ruthlessly with the same brand on each microscope. A lot of our live cell work is/was done with low power [20x Phase] air objectives though.
>
> I do notice that immersion oil often ingresses into oil immersion objective internal optics with time, although strangely it rarely notices when imaging [like dirt on much of the internal optics, you must mostly focus through it]. One microscopy core manager even commented to me that he considers oil objectives ‘consumables’. The cost of ‘repair’ [often the same price as a new objective] is so high we frequently have to live with it [I advise users that our say 40x objective image quality is suspect for this reason]. In truth we don’t notice any drop in image quality when it must have happened, although you can see it ‘smeared’ on the optics when looking through the removed objective, using a magnifying glass, bright room light and a ‘cleaned’ objective top lens.
>
> Old listerver posting on Cargille 37 attached below.
>
> Regards
>
> Keith
>
> PS missed this link on the last post:
>
> Dissolving crystals in Zeiss 518F oil
> http://www.zeiss.com/C1256F8500454979/0/E8CD09DFA520787FC1256F86004B5FFB/$file/immersol-crystals.pdf
>
> ---------------------------------------------------------------------------
> Dr Keith J. Morris,
> Molecular Cytogenetics and Microscopy Core,
> Laboratory 00/069 and 00/070,
> The Wellcome Trust Centre for Human Genetics,
> Roosevelt Drive,
> Oxford  OX3 7BN,
> United Kingdom.
>
> Telephone:  [phone]
> Email:  [hidden email]
> Web-pages: http://www.well.ox.ac.uk/cytogenetics/
>
>
>
> Subject:
>
> Re: Fluorescence from Type 37 Immersion Oil
>
> From:
>
> Vitaly Boyko <[hidden email]>
>
> Reply-To:
>
> Vitaly Boyko <[hidden email]>
>
> Date:
>
> Wed, 16 Aug 2006 10:23:20 -0400
>
> Content-Type:
>
> text/plain
>
>
>
> Search the CONFOCAL archive at
>
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
>
>
> PS What would be more appropriate to compare is Zeiss 518F (not 518C!) with
>
> the Nikon type A or Cargille type DF.
>
>
>
>
>
>
>
> Hi Jason,
>
>
>
> I wonder why you have tested the Cargille FF imm. oil, as the refractive
>
> index of FF oil is too far off for the standard glass (1.518) ?????!!!!!
>
>
>
> Actually, I haven't noticed the "very bad" autofluorescence of the Cargille
>
> type DF versus Zeiss 518 in the CFP, YFP or FRET channels. The difference is
>
> less than 10%.
>
>
>
> The problem with the Nikon oil is its dispenser and/or viscosity - it is
>
> often annoying "fighting" with the air bubbles.
>
>
>
> What is about the Invitrogen/Mol.Probes mounting media autofluorescence?
>
>
>
> I am fed up with the irreproducibility of the background autofluorescence
>
> using the ProlongGold mounting media!!! That is a serious problem compared
>
> to the issue of the immersion oils.
>
>
>
> Cheers,
>
>
>
> Vitaly
>
>
>
> NCI-Frederick
>
>
>
>
>
> ----- Original Message -----
>
> From: "Kilgore, Jason" <[hidden email]>
>
> To: <[hidden email]>
>
> Sent: Tuesday, August 15, 2006 7:26 PM
>
> Subject: Re: Fluorescence from Type 37 Immersion Oil
>
>
>
>
>
> > Search the CONFOCAL archive at
>
> > http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
> >
>
> > Back in 2000 I performed a comparison of four different immersion oils
>
> > (Zeiss 518C, Nikon Type A, Cargille Type DF, and Cargille Type FF) for
>
> > autofluorescence (using DAPI, FITC, acridine orange, and TRITC filter
>
> > sets) and general resolution at 60x on a Nikon E400.  I didn't test
>
> > Cargille 37.
>
> >
>
> > Green autofluorescence tended to be worst for all four, but was least
>
> > for Cargille FF, followed by Nikon Type A, then Zeiss, then (very bad)
>
> > Cargille DF.  However, the resolution was unacceptable for Cargille FF.
>
> > Autofluorescence in the other channels was moderate (DAPI, TRITC) to
>
> > very low (acridine orange), with the same comparative trend applying.
>
> >
>
> > Thus, ever since we have been using Nikon Type A with no complaints for
>
> > most of our imaging.
>
> >
>
> > Feel free to contact me personally and I can send you the data.
>
> >
>
> > Jason
>
> >
>
> > Jason A. Kilgore
>
> > Cell Biology / Histology
>
> > Molecular Probes/Invitrogen
>
> > [hidden email]
>
> >
>
> >
>
> > -----Original Message-----
>
> > From: Confocal Microscopy List [mailto:[hidden email]] On
>
> > Behalf Of S. Brunet
>
> > Sent: Tuesday, August 15, 2006 11:19 AM
>
> > To: [hidden email]
>
> > Subject: Fluorescence from Type 37 Immersion Oil
>
> >
>
> > Search the CONFOCAL archive at
>
> > http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
> >
>
> > Hello to all,
>
> >
>
> > We recently began using the Cargille 37 oil with the temperature chamber
>
> > on the
>
> > microscope.  I found significant fluorescence from the oil when exposed
>
> > to the
>
> > HBO lamp (I could not see the eYFP because of the blue background).
>
> >
>
> > So I used the 20x air objective and I was able to obtain an image from
>
> > the
>
> > fluorescence from a drop of 37 oil placed on a coverslip as a sample
>
> > (excitation at 488 or 458nm and pinhole at the usual confocal setting).
>
> >
>
> > Is this the norm?
>
> >
>
> > Regards,
>
> > Sophie
>
> > _______________________________________
>
> > Sophie M.K. Brunet, Ph.D.
>
> > Research Officer
>
> > Optical Spectroscopist, Laser Systems and Applications
>
> > [hidden email]
>
> > 306-966-1719 (office)   306-966-1702 (fax)
>
> > _______________________________________
>
> > Saskatchewan Structural Sciences Centre
>
> > University of Saskatchewan
>
> > Thorvaldson Bldg.
>
> > 110 Science Place
>
> > Saskatoon, SK   S7N 5C9
>
> >
>
>

[John Oreopoulos]

We purchased a bottle of this product a few months back and I've used  
it a few times when doing live-cell imaging. One thing I noticed that  
was different when using this compared to regular immersion oil for  
fluorescence was that there was a short lag time in terms of  
adjusting the focus of the microscope and the image coming into focus  
on my preview computer screen (and in the eyepieces). I guess this  
has to do with the viscosity of the oil. It took a while to get  
adjusted to this because I would keep overshooting the fine focus,  
but other than that it works great (no commercial interest).

John Oreopoulos


On 23-Mar-09, at 6:42 AM, Christophe Leterrier wrote:

> I would like to point out that Cargille has a new oil for fluorescence
> imaging at 37°C, called 37DF, that has very low auto fluorescence. It
> is not on their website, but I could get the product pdf sent to me by
> a Cargille person, so I can send it if someone is interested. I'm
> trying to get some here in France, but none of the french Cargille
> distributors has heard about it yet... It has been found to be quite
> good for fluorescence imaging of live cells at 37°C, according to this
> paper in J Microscopy :
>
> Immersion oil for high-resolution live-cell imaging at 37 degrees C:
> optical and physical characteristics.
> Oomen LC, Sacher R, Brocks HH, Zwier JM, Brakenhoff GJ, Jalink K.
> J Microsc. 2008 Nov;232(2):353-61.
>
>
> On Mon, Mar 23, 2009 at 11:18, Keith Morris  
> <[hidden email]> wrote:
>>
>> Hi all,
>>
>>
>>
>> Using immersion oil at 37oC.
>>
>>
>>
>> Zeiss sell a range of low fluorescence immersion oils: bottles of  
>> 20, 250 and 500 ml, unfortunately all filled with the same 518F  
>> oil. According to Zeiss it’s apparently ‘compatible’ with our  
>> Zeiss microscopes, but little extra information is offered on the  
>> Zeiss website.
>>
>>
>>
>> The only info I can gleam from the web [other than the ubiquitous  
>> 518F safety data Sheet] is the following
>>
>> Immersion oil "Immersol" 518 F fluorescence free, (ISO 8036-1/2, n
>> (e)=1.518 (23°C), halogen-free)
>>
>>
>>
>> So not much comment about 37oC use.  PeCon’s site [who supply most  
>> of Zeiss’s decent incubator stuff] have no opinion on the matter  
>> either.
>>
>>
>>
>> The 518F oil safety sheet mentions “Use of the Substance /  
>> Preparation: For application information, consult the processing  
>> instructions. Processing instructions or technical
>>
>> information sheet available on request.”. I’ll ask our  
>> knowledgeable local Zeiss Confocal rep for more details, as we are  
>> just installing a Zeiss/PeCon 37oC live cell incubator here on our  
>> Zeiss 510 MetaHead confocal.
>>
>> Cargilee’s site in comparison is passionate about all things  
>> immersion oil:
>> http://www.cargille.com/immeroil.shtml
>> http://www.cargille.com/immeroilselection.shtml
>> and their excellent, if elderly, ‘primer’
>> http://www.cargille.com/immersionoilmicroscope.shtml
>>
>> From postings to this server back in 2006, and Cargille’s site not  
>> mentioning ‘non-fluorescence’ for type 37, it appears that  
>> Cargille 37 probably autofluoresces, so it’s likely better for  
>> live cell phase-contrast [halogen lamp] transmission microscopy  
>> [but then often so are dry objectives]. I’ll stick with standard  
>> 518F immersion oil at 37oC though, it’s always imaged well enough  
>> on Zeiss microscopes at that temperature [and at least at 37oC it  
>> never became cloudy due to crystal precipitation]. Similarly the  
>> Cargille DF and FF seem fine to me at 37oC, and we stick  
>> ruthlessly with the same brand on each microscope. A lot of our  
>> live cell work is/was done with low power [20x Phase] air  
>> objectives though.
>>
>> I do notice that immersion oil often ingresses into oil immersion  
>> objective internal optics with time, although strangely it rarely  
>> notices when imaging [like dirt on much of the internal optics,  
>> you must mostly focus through it]. One microscopy core manager  
>> even commented to me that he considers oil objectives  
>> ‘consumables’. The cost of ‘repair’ [often the same price as a new  
>> objective] is so high we frequently have to live with it [I advise  
>> users that our say 40x objective image quality is suspect for this  
>> reason]. In truth we don’t notice any drop in image quality when  
>> it must have happened, although you can see it ‘smeared’ on the  
>> optics when looking through the removed objective, using a  
>> magnifying glass, bright room light and a ‘cleaned’ objective top  
>> lens.
>>
>> Old listerver posting on Cargille 37 attached below.
>>
>> Regards
>>
>> Keith
>>
>> PS missed this link on the last post:
>>
>> Dissolving crystals in Zeiss 518F oil
>> http://www.zeiss.com/C1256F8500454979/0/ 
>> E8CD09DFA520787FC1256F86004B5FFB/$file/immersol-crystals.pdf
>>
>> ---------------------------------------------------------------------
>> ------
>> Dr Keith J. Morris,
>> Molecular Cytogenetics and Microscopy Core,
>> Laboratory 00/069 and 00/070,
>> The Wellcome Trust Centre for Human Genetics,
>> Roosevelt Drive,
>> Oxford  OX3 7BN,
>> United Kingdom.
>>
>> Telephone:  [phone]
>> Email:  [hidden email]
>> Web-pages: http://www.well.ox.ac.uk/cytogenetics/
>>
>>
>>
>> Subject:
>>
>> Re: Fluorescence from Type 37 Immersion Oil
>>
>> From:
>>
>> Vitaly Boyko <[hidden email]>
>>
>> Reply-To:
>>
>> Vitaly Boyko <[hidden email]>
>>
>> Date:
>>
>> Wed, 16 Aug 2006 10:23:20 -0400
>>
>> Content-Type:
>>
>> text/plain
>>
>>
>>
>> Search the CONFOCAL archive at
>>
>> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>>
>>
>>
>> PS What would be more appropriate to compare is Zeiss 518F (not  
>> 518C!) with
>>
>> the Nikon type A or Cargille type DF.
>>
>>
>>
>>
>>
>>
>>
>> Hi Jason,
>>
>>
>>
>> I wonder why you have tested the Cargille FF imm. oil, as the  
>> refractive
>>
>> index of FF oil is too far off for the standard glass  
>> (1.518) ?????!!!!!
>>
>>
>>
>> Actually, I haven't noticed the "very bad" autofluorescence of the  
>> Cargille
>>
>> type DF versus Zeiss 518 in the CFP, YFP or FRET channels. The  
>> difference is
>>
>> less than 10%.
>>
>>
>>
>> The problem with the Nikon oil is its dispenser and/or viscosity -  
>> it is
>>
>> often annoying "fighting" with the air bubbles.
>>
>>
>>
>> What is about the Invitrogen/Mol.Probes mounting media  
>> autofluorescence?
>>
>>
>>
>> I am fed up with the irreproducibility of the background  
>> autofluorescence
>>
>> using the ProlongGold mounting media!!! That is a serious problem  
>> compared
>>
>> to the issue of the immersion oils.
>>
>>
>>
>> Cheers,
>>
>>
>>
>> Vitaly
>>
>>
>>
>> NCI-Frederick
>>
>>
>>
>>
>>
>> ----- Original Message -----
>>
>> From: "Kilgore, Jason" <[hidden email]>
>>
>> To: <[hidden email]>
>>
>> Sent: Tuesday, August 15, 2006 7:26 PM
>>
>> Subject: Re: Fluorescence from Type 37 Immersion Oil
>>
>>
>>
>>
>>
>>> Search the CONFOCAL archive at
>>
>>> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>>
>>>
>>
>>> Back in 2000 I performed a comparison of four different immersion  
>>> oils
>>
>>> (Zeiss 518C, Nikon Type A, Cargille Type DF, and Cargille Type  
>>> FF) for
>>
>>> autofluorescence (using DAPI, FITC, acridine orange, and TRITC  
>>> filter
>>
>>> sets) and general resolution at 60x on a Nikon E400.  I didn't test
>>
>>> Cargille 37.
>>
>>>
>>
>>> Green autofluorescence tended to be worst for all four, but was  
>>> least
>>
>>> for Cargille FF, followed by Nikon Type A, then Zeiss, then (very  
>>> bad)
>>
>>> Cargille DF.  However, the resolution was unacceptable for  
>>> Cargille FF.
>>
>>> Autofluorescence in the other channels was moderate (DAPI, TRITC) to
>>
>>> very low (acridine orange), with the same comparative trend  
>>> applying.
>>
>>>
>>
>>> Thus, ever since we have been using Nikon Type A with no  
>>> complaints for
>>
>>> most of our imaging.
>>
>>>
>>
>>> Feel free to contact me personally and I can send you the data.
>>
>>>
>>
>>> Jason
>>
>>>
>>
>>> Jason A. Kilgore
>>
>>> Cell Biology / Histology
>>
>>> Molecular Probes/Invitrogen
>>
>>> [hidden email]
>>
>>>
>>
>>>
>>
>>> -----Original Message-----
>>
>>> From: Confocal Microscopy List  
>>> [mailto:[hidden email]] On
>>
>>> Behalf Of S. Brunet
>>
>>> Sent: Tuesday, August 15, 2006 11:19 AM
>>
>>> To: [hidden email]
>>
>>> Subject: Fluorescence from Type 37 Immersion Oil
>>
>>>
>>
>>> Search the CONFOCAL archive at
>>
>>> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>>
>>>
>>
>>> Hello to all,
>>
>>>
>>
>>> We recently began using the Cargille 37 oil with the temperature  
>>> chamber
>>
>>> on the
>>
>>> microscope.  I found significant fluorescence from the oil when  
>>> exposed
>>
>>> to the
>>
>>> HBO lamp (I could not see the eYFP because of the blue background).
>>
>>>
>>
>>> So I used the 20x air objective and I was able to obtain an image  
>>> from
>>
>>> the
>>
>>> fluorescence from a drop of 37 oil placed on a coverslip as a sample
>>
>>> (excitation at 488 or 458nm and pinhole at the usual confocal  
>>> setting).
>>
>>>
>>
>>> Is this the norm?
>>
>>>
>>
>>> Regards,
>>
>>> Sophie
>>
>>> _______________________________________
>>
>>> Sophie M.K. Brunet, Ph.D.
>>
>>> Research Officer
>>
>>> Optical Spectroscopist, Laser Systems and Applications
>>
>>> [hidden email]
>>
>>> 306-966-1719 (office)   306-966-1702 (fax)
>>
>>> _______________________________________
>>
>>> Saskatchewan Structural Sciences Centre
>>
>>> University of Saskatchewan
>>
>>> Thorvaldson Bldg.
>>
>>> 110 Science Place
>>
>>> Saskatoon, SK   S7N 5C9
>>
>>>
>>
>>

[Lauran Oomen]

Dear all,

As someone who is so directly involved (being the first author of the publication mentioned below), at first I resisted reacting to this thread. Now I would like to mention that information about the 37DF immersion oil can be found on Cargille's website (http://www.cargille.com/immeroil.shtml) and secondly, that those of you who might be interested in a pdf of our publication on this oil can write directly to me.

Please note: NO commercial interest.

Lauran
--------------------------------------------------------------
Lauran Oomen
Manager Digital Microscopy Facility (H003)
NKI-AVL
Plesmanlaan 121
PO Box 90203
1006 BE Amsterdam
The Netherlands
 
phone [phone]
[hidden email]

-----Original Message-----
From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Christophe Leterrier
Sent: maandag 23 maart 2009 11:42
To: [hidden email]
Subject: Re: Immersion Oil - 37oC

I would like to point out that Cargille has a new oil for fluorescence
imaging at 37°C, called 37DF, that has very low auto fluorescence. It
is not on their website, but I could get the product pdf sent to me by
a Cargille person, so I can send it if someone is interested. I'm
trying to get some here in France, but none of the french Cargille
distributors has heard about it yet... It has been found to be quite
good for fluorescence imaging of live cells at 37°C, according to this
paper in J Microscopy :

Immersion oil for high-resolution live-cell imaging at 37 degrees C:
optical and physical characteristics.
Oomen LC, Sacher R, Brocks HH, Zwier JM, Brakenhoff GJ, Jalink K.
J Microsc. 2008 Nov;232(2):353-61.


On Mon, Mar 23, 2009 at 11:18, Keith Morris <[hidden email]> wrote:

>
> Hi all,
>
>
>
> Using immersion oil at 37oC.
>
>
>
> Zeiss sell a range of low fluorescence immersion oils: bottles of 20, 250 and 500 ml, unfortunately all filled with the same 518F oil. According to Zeiss it's apparently 'compatible' with our Zeiss microscopes, but little extra information is offered on the Zeiss website.
>
>
>
> The only info I can gleam from the web [other than the ubiquitous 518F safety data Sheet] is the following
>
> Immersion oil "Immersol" 518 F fluorescence free, (ISO 8036-1/2, n(e)=1.518 (23°C), halogen-free)
>
>
>
> So not much comment about 37oC use.  PeCon's site [who supply most of Zeiss's decent incubator stuff] have no opinion on the matter either.
>
>
>
> The 518F oil safety sheet mentions "Use of the Substance / Preparation: For application information, consult the processing instructions. Processing instructions or technical
>
> information sheet available on request.". I'll ask our knowledgeable local Zeiss Confocal rep for more details, as we are just installing a Zeiss/PeCon 37oC live cell incubator here on our Zeiss 510 MetaHead confocal.
>
> Cargilee's site in comparison is passionate about all things immersion oil:
> http://www.cargille.com/immeroil.shtml
> http://www.cargille.com/immeroilselection.shtml
> and their excellent, if elderly, 'primer'
> http://www.cargille.com/immersionoilmicroscope.shtml
>
> From postings to this server back in 2006, and Cargille's site not mentioning 'non-fluorescence' for type 37, it appears that Cargille 37 probably autofluoresces, so it's likely better for live cell phase-contrast [halogen lamp] transmission microscopy [but then often so are dry objectives]. I'll stick with standard 518F immersion oil at 37oC though, it's always imaged well enough on Zeiss microscopes at that temperature [and at least at 37oC it never became cloudy due to crystal precipitation]. Similarly the Cargille DF and FF seem fine to me at 37oC, and we stick ruthlessly with the same brand on each microscope. A lot of our live cell work is/was done with low power [20x Phase] air objectives though.
>
> I do notice that immersion oil often ingresses into oil immersion objective internal optics with time, although strangely it rarely notices when imaging [like dirt on much of the internal optics, you must mostly focus through it]. One microscopy core manager even commented to me that he considers oil objectives 'consumables'. The cost of 'repair' [often the same price as a new objective] is so high we frequently have to live with it [I advise users that our say 40x objective image quality is suspect for this reason]. In truth we don't notice any drop in image quality when it must have happened, although you can see it 'smeared' on the optics when looking through the removed objective, using a magnifying glass, bright room light and a 'cleaned' objective top lens.
>
> Old listerver posting on Cargille 37 attached below.
>
> Regards
>
> Keith
>
> PS missed this link on the last post:
>
> Dissolving crystals in Zeiss 518F oil
> http://www.zeiss.com/C1256F8500454979/0/E8CD09DFA520787FC1256F86004B5FFB/$file/immersol-crystals.pdf
>
> ---------------------------------------------------------------------------
> Dr Keith J. Morris,
> Molecular Cytogenetics and Microscopy Core,
> Laboratory 00/069 and 00/070,
> The Wellcome Trust Centre for Human Genetics,
> Roosevelt Drive,
> Oxford  OX3 7BN,
> United Kingdom.
>
> Telephone:  [phone]
> Email:  [hidden email]
> Web-pages: http://www.well.ox.ac.uk/cytogenetics/
>
>
>
> Subject:
>
> Re: Fluorescence from Type 37 Immersion Oil
>
> From:
>
> Vitaly Boyko <[hidden email]>
>
> Reply-To:
>
> Vitaly Boyko <[hidden email]>
>
> Date:
>
> Wed, 16 Aug 2006 10:23:20 -0400
>
> Content-Type:
>
> text/plain
>
>
>
> Search the CONFOCAL archive at
>
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
>
>
> PS What would be more appropriate to compare is Zeiss 518F (not 518C!) with
>
> the Nikon type A or Cargille type DF.
>
>
>
>
>
>
>
> Hi Jason,
>
>
>
> I wonder why you have tested the Cargille FF imm. oil, as the refractive
>
> index of FF oil is too far off for the standard glass (1.518) ?????!!!!!
>
>
>
> Actually, I haven't noticed the "very bad" autofluorescence of the Cargille
>
> type DF versus Zeiss 518 in the CFP, YFP or FRET channels. The difference is
>
> less than 10%.
>
>
>
> The problem with the Nikon oil is its dispenser and/or viscosity - it is
>
> often annoying "fighting" with the air bubbles.
>
>
>
> What is about the Invitrogen/Mol.Probes mounting media autofluorescence?
>
>
>
> I am fed up with the irreproducibility of the background autofluorescence
>
> using the ProlongGold mounting media!!! That is a serious problem compared
>
> to the issue of the immersion oils.
>
>
>
> Cheers,
>
>
>
> Vitaly
>
>
>
> NCI-Frederick
>
>
>
>
>
> ----- Original Message -----
>
> From: "Kilgore, Jason" <[hidden email]>
>
> To: <[hidden email]>
>
> Sent: Tuesday, August 15, 2006 7:26 PM
>
> Subject: Re: Fluorescence from Type 37 Immersion Oil
>
>
>
>
>
> > Search the CONFOCAL archive at
>
> > http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
> >
>
> > Back in 2000 I performed a comparison of four different immersion oils
>
> > (Zeiss 518C, Nikon Type A, Cargille Type DF, and Cargille Type FF) for
>
> > autofluorescence (using DAPI, FITC, acridine orange, and TRITC filter
>
> > sets) and general resolution at 60x on a Nikon E400.  I didn't test
>
> > Cargille 37.
>
> >
>
> > Green autofluorescence tended to be worst for all four, but was least
>
> > for Cargille FF, followed by Nikon Type A, then Zeiss, then (very bad)
>
> > Cargille DF.  However, the resolution was unacceptable for Cargille FF.
>
> > Autofluorescence in the other channels was moderate (DAPI, TRITC) to
>
> > very low (acridine orange), with the same comparative trend applying.
>
> >
>
> > Thus, ever since we have been using Nikon Type A with no complaints for
>
> > most of our imaging.
>
> >
>
> > Feel free to contact me personally and I can send you the data.
>
> >
>
> > Jason
>
> >
>
> > Jason A. Kilgore
>
> > Cell Biology / Histology
>
> > Molecular Probes/Invitrogen
>
> > [hidden email]
>
> >
>
> >
>
> > -----Original Message-----
>
> > From: Confocal Microscopy List [mailto:[hidden email]] On
>
> > Behalf Of S. Brunet
>
> > Sent: Tuesday, August 15, 2006 11:19 AM
>
> > To: [hidden email]
>
> > Subject: Fluorescence from Type 37 Immersion Oil
>
> >
>
> > Search the CONFOCAL archive at
>
> > http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
> >
>
> > Hello to all,
>
> >
>
> > We recently began using the Cargille 37 oil with the temperature chamber
>
> > on the
>
> > microscope.  I found significant fluorescence from the oil when exposed
>
> > to the
>
> > HBO lamp (I could not see the eYFP because of the blue background).
>
> >
>
> > So I used the 20x air objective and I was able to obtain an image from
>
> > the
>
> > fluorescence from a drop of 37 oil placed on a coverslip as a sample
>
> > (excitation at 488 or 458nm and pinhole at the usual confocal setting).
>
> >
>
> > Is this the norm?
>
> >
>
> > Regards,
>
> > Sophie
>
> > _______________________________________
>
> > Sophie M.K. Brunet, Ph.D.
>
> > Research Officer
>
> > Optical Spectroscopist, Laser Systems and Applications
>
> > [hidden email]
>
> > 306-966-1719 (office)   306-966-1702 (fax)
>
> > _______________________________________
>
> > Saskatchewan Structural Sciences Centre
>
> > University of Saskatchewan
>
> > Thorvaldson Bldg.
>
> > 110 Science Place
>
> > Saskatoon, SK   S7N 5C9
>
> >
>
>

[Keith Morris]

The 37DF must have just arrived on the website [and there's no spec on it as
I write], Thanks for pointing out its existence.

As we will continue to use Zeiss 518F at low temperature on our Zeiss
microscopes we won't change to a Cargille oil for 37oC, but it’s interesting
to know one exists optimised for 37oC and fluorescence microscopy. In
practice as live cells are moving objects you will possibly take a hit in
image absolute quality at high mag anyway when imaging fluorescence at 37oC.
For those using Cargille oils at low temps you have the real advantage given
Cargille oils are all touted as miscible with each other.  

Keith  

---------------------------------------------------------------------------
Dr Keith J. Morris,
Molecular Cytogenetics and Microscopy Core,
Laboratory 00/069 and 00/070,
The Wellcome Trust Centre for Human Genetics,
Roosevelt Drive,
Oxford  OX3 7BN,
United Kingdom.

Telephone:  [phone]
Email:  [hidden email]
Web-pages: http://www.well.ox.ac.uk/cytogenetics/
 

-----Original Message-----
From: Confocal Microscopy List [mailto:[hidden email]] On
Behalf Of John Oreopoulos
Sent: 23 March 2009 12:56
To: [hidden email]
Subject: Re: Immersion Oil - 37oC

We purchased a bottle of this product a few months back and I've used  
it a few times when doing live-cell imaging. One thing I noticed that  
was different when using this compared to regular immersion oil for  
fluorescence was that there was a short lag time in terms of  
adjusting the focus of the microscope and the image coming into focus  
on my preview computer screen (and in the eyepieces). I guess this  
has to do with the viscosity of the oil. It took a while to get  
adjusted to this because I would keep overshooting the fine focus,  
but other than that it works great (no commercial interest).

John Oreopoulos


On 23-Mar-09, at 6:42 AM, Christophe Leterrier wrote:

> I would like to point out that Cargille has a new oil for fluorescence
> imaging at 37°C, called 37DF, that has very low auto fluorescence. It
> is not on their website, but I could get the product pdf sent to me by
> a Cargille person, so I can send it if someone is interested. I'm
> trying to get some here in France, but none of the french Cargille
> distributors has heard about it yet... It has been found to be quite
> good for fluorescence imaging of live cells at 37°C, according to this
> paper in J Microscopy :
>
> Immersion oil for high-resolution live-cell imaging at 37 degrees C:
> optical and physical characteristics.
> Oomen LC, Sacher R, Brocks HH, Zwier JM, Brakenhoff GJ, Jalink K.
> J Microsc. 2008 Nov;232(2):353-61.
>
>
> On Mon, Mar 23, 2009 at 11:18, Keith Morris  
> <[hidden email]> wrote:
>>
>> Hi all,
>>
>>
>>
>> Using immersion oil at 37oC.
>>
>>
>>
>> Zeiss sell a range of low fluorescence immersion oils: bottles of  
>> 20, 250 and 500 ml, unfortunately all filled with the same 518F  
>> oil. According to Zeiss it’s apparently ‘compatible’ with our  
>> Zeiss microscopes, but little extra information is offered on the  
>> Zeiss website.
>>
>>
>>
>> The only info I can gleam from the web [other than the ubiquitous  
>> 518F safety data Sheet] is the following
>>
>> Immersion oil "Immersol" 518 F fluorescence free, (ISO 8036-1/2, n
>> (e)=1.518 (23°C), halogen-free)
>>
>>
>>
>> So not much comment about 37oC use.  PeCon’s site [who supply most  
>> of Zeiss’s decent incubator stuff] have no opinion on the matter  
>> either.
>>
>>
>>
>> The 518F oil safety sheet mentions “Use of the Substance /  
>> Preparation: For application information, consult the processing  
>> instructions. Processing instructions or technical
>>
>> information sheet available on request.”. I’ll ask our  
>> knowledgeable local Zeiss Confocal rep for more details, as we are  
>> just installing a Zeiss/PeCon 37oC live cell incubator here on our  
>> Zeiss 510 MetaHead confocal.
>>
>> Cargilee’s site in comparison is passionate about all things  
>> immersion oil:
>> http://www.cargille.com/immeroil.shtml
>> http://www.cargille.com/immeroilselection.shtml
>> and their excellent, if elderly, ‘primer’
>> http://www.cargille.com/immersionoilmicroscope.shtml
>>
>> From postings to this server back in 2006, and Cargille’s site not  
>> mentioning ‘non-fluorescence’ for type 37, it appears that  
>> Cargille 37 probably autofluoresces, so it’s likely better for  
>> live cell phase-contrast [halogen lamp] transmission microscopy  
>> [but then often so are dry objectives]. I’ll stick with standard  
>> 518F immersion oil at 37oC though, it’s always imaged well enough  
>> on Zeiss microscopes at that temperature [and at least at 37oC it  
>> never became cloudy due to crystal precipitation]. Similarly the  
>> Cargille DF and FF seem fine to me at 37oC, and we stick  
>> ruthlessly with the same brand on each microscope. A lot of our  
>> live cell work is/was done with low power [20x Phase] air  
>> objectives though.
>>
>> I do notice that immersion oil often ingresses into oil immersion  
>> objective internal optics with time, although strangely it rarely  
>> notices when imaging [like dirt on much of the internal optics,  
>> you must mostly focus through it]. One microscopy core manager  
>> even commented to me that he considers oil objectives  
>> ‘consumables’. The cost of ‘repair’ [often the same price as a new  
>> objective] is so high we frequently have to live with it [I advise  
>> users that our say 40x objective image quality is suspect for this  
>> reason]. In truth we don’t notice any drop in image quality when  
>> it must have happened, although you can see it ‘smeared’ on the  
>> optics when looking through the removed objective, using a  
>> magnifying glass, bright room light and a ‘cleaned’ objective top  
>> lens.
>>
>> Old listerver posting on Cargille 37 attached below.
>>
>> Regards
>>
>> Keith
>>
>> PS missed this link on the last post:
>>
>> Dissolving crystals in Zeiss 518F oil
>> http://www.zeiss.com/C1256F8500454979/0/ 
>> E8CD09DFA520787FC1256F86004B5FFB/$file/immersol-crystals.pdf
>>
>> ---------------------------------------------------------------------
>> ------
>> Dr Keith J. Morris,
>> Molecular Cytogenetics and Microscopy Core,
>> Laboratory 00/069 and 00/070,
>> The Wellcome Trust Centre for Human Genetics,
>> Roosevelt Drive,
>> Oxford  OX3 7BN,
>> United Kingdom.
>>
>> Telephone:  [phone]
>> Email:  [hidden email]
>> Web-pages: http://www.well.ox.ac.uk/cytogenetics/
>>
>>
>>
>> Subject:
>>
>> Re: Fluorescence from Type 37 Immersion Oil
>>
>> From:
>>
>> Vitaly Boyko <[hidden email]>
>>
>> Reply-To:
>>
>> Vitaly Boyko <[hidden email]>
>>
>> Date:
>>
>> Wed, 16 Aug 2006 10:23:20 -0400
>>
>> Content-Type:
>>
>> text/plain
>>
>>
>>
>> Search the CONFOCAL archive at
>>
>> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>>
>>
>>
>> PS What would be more appropriate to compare is Zeiss 518F (not  
>> 518C!) with
>>
>> the Nikon type A or Cargille type DF.
>>
>>
>>
>>
>>
>>
>>
>> Hi Jason,
>>
>>
>>
>> I wonder why you have tested the Cargille FF imm. oil, as the  
>> refractive
>>
>> index of FF oil is too far off for the standard glass  
>> (1.518) ?????!!!!!
>>
>>
>>
>> Actually, I haven't noticed the "very bad" autofluorescence of the  
>> Cargille
>>
>> type DF versus Zeiss 518 in the CFP, YFP or FRET channels. The  
>> difference is
>>
>> less than 10%.
>>
>>
>>
>> The problem with the Nikon oil is its dispenser and/or viscosity -  
>> it is
>>
>> often annoying "fighting" with the air bubbles.
>>
>>
>>
>> What is about the Invitrogen/Mol.Probes mounting media  
>> autofluorescence?
>>
>>
>>
>> I am fed up with the irreproducibility of the background  
>> autofluorescence
>>
>> using the ProlongGold mounting media!!! That is a serious problem  
>> compared
>>
>> to the issue of the immersion oils.
>>
>>
>>
>> Cheers,
>>
>>
>>
>> Vitaly
>>
>>
>>
>> NCI-Frederick
>>
>>
>>
>>
>>
>> ----- Original Message -----
>>
>> From: "Kilgore, Jason" <[hidden email]>
>>
>> To: <[hidden email]>
>>
>> Sent: Tuesday, August 15, 2006 7:26 PM
>>
>> Subject: Re: Fluorescence from Type 37 Immersion Oil
>>
>>
>>
>>
>>
>>> Search the CONFOCAL archive at
>>
>>> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>>
>>>
>>
>>> Back in 2000 I performed a comparison of four different immersion  
>>> oils
>>
>>> (Zeiss 518C, Nikon Type A, Cargille Type DF, and Cargille Type  
>>> FF) for
>>
>>> autofluorescence (using DAPI, FITC, acridine orange, and TRITC  
>>> filter
>>
>>> sets) and general resolution at 60x on a Nikon E400.  I didn't test
>>
>>> Cargille 37.
>>
>>>
>>
>>> Green autofluorescence tended to be worst for all four, but was  
>>> least
>>
>>> for Cargille FF, followed by Nikon Type A, then Zeiss, then (very  
>>> bad)
>>
>>> Cargille DF.  However, the resolution was unacceptable for  
>>> Cargille FF.
>>
>>> Autofluorescence in the other channels was moderate (DAPI, TRITC) to
>>
>>> very low (acridine orange), with the same comparative trend  
>>> applying.
>>
>>>
>>
>>> Thus, ever since we have been using Nikon Type A with no  
>>> complaints for
>>
>>> most of our imaging.
>>
>>>
>>
>>> Feel free to contact me personally and I can send you the data.
>>
>>>
>>
>>> Jason
>>
>>>
>>
>>> Jason A. Kilgore
>>
>>> Cell Biology / Histology
>>
>>> Molecular Probes/Invitrogen
>>
>>> [hidden email]
>>
>>>
>>
>>>
>>
>>> -----Original Message-----
>>
>>> From: Confocal Microscopy List  
>>> [mailto:[hidden email]] On
>>
>>> Behalf Of S. Brunet
>>
>>> Sent: Tuesday, August 15, 2006 11:19 AM
>>
>>> To: [hidden email]
>>
>>> Subject: Fluorescence from Type 37 Immersion Oil
>>
>>>
>>
>>> Search the CONFOCAL archive at
>>
>>> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>>
>>>
>>
>>> Hello to all,
>>
>>>
>>
>>> We recently began using the Cargille 37 oil with the temperature  
>>> chamber
>>
>>> on the
>>
>>> microscope.  I found significant fluorescence from the oil when  
>>> exposed
>>
>>> to the
>>
>>> HBO lamp (I could not see the eYFP because of the blue background).
>>
>>>
>>
>>> So I used the 20x air objective and I was able to obtain an image  
>>> from
>>
>>> the
>>
>>> fluorescence from a drop of 37 oil placed on a coverslip as a sample
>>
>>> (excitation at 488 or 458nm and pinhole at the usual confocal  
>>> setting).
>>
>>>
>>
>>> Is this the norm?
>>
>>>
>>
>>> Regards,
>>
>>> Sophie
>>
>>> _______________________________________
>>
>>> Sophie M.K. Brunet, Ph.D.
>>
>>> Research Officer
>>
>>> Optical Spectroscopist, Laser Systems and Applications
>>
>>> [hidden email]
>>
>>> 306-966-1719 (office)   306-966-1702 (fax)
>>
>>> _______________________________________
>>
>>> Saskatchewan Structural Sciences Centre
>>
>>> University of Saskatchewan
>>
>>> Thorvaldson Bldg.
>>
>>> 110 Science Place
>>
>>> Saskatoon, SK   S7N 5C9
>>
>>>
>>
>>

[Keith Morris]

Back in 2006 I wrote the following to the microscopy.com listerver in
response to a query about the acidity [pH] of immersion oil. As it seems
relevant to this immersion oil thread I'll repost it here for those with a
passing interest [and it mention's Cargille DF's old warning label from 4+
years ago - in 2009, Cargille DF bottles carry no such warning]:
---------------------------------------------------------------------------
Hi,

Question:

What is the pH of a cedarwood oilJuniperus virginiana) for oil
immersion objective? what is the pH of lemongrass oil(Cymbopogon citratus)?

Reply:

I was under the impression that, although oils can contain acidic
contaminates, these are often removed during the refining process (not being
good for engines etc.). Acidity (the acid-type constituents) "is usually
defined in terms of total acid number (TAN). These constituents vary in
nature and may or may not markedly influence the behaviour of the
lubricant". Low acidity oils can oxidise during use and become more acidic
over time (years) - e.g. the acid number goes to over 0.4 - while the newly
refined 'non-corrosive' oil would be expected to be nearer to 0.05. The
acids can be weak organic and strong inorganic. Acidity in oil can be useful
for things like improved water resistance. Oils don't have an aqueous pH as
such though, just the TAN number (total acidic number).

Olive oil for example is also graded for consumption on it's oleic acid
acidity - around 3% in cheaper olive oil, and down to below 1% in 'extra
virgin'. Oils are generally insoluble in water, which rather helps reduce
it's corrosive properties during use.

Cargille use the Neutralization Number for their immersion oils. This is a
measure of the acidity or alkalinity of the oil and is the mass in
milligrams of the amount of acid (HCl) or base (KOH) required to neutralize
one gram of the oil. For their standard immersion oils and type FF the value
is very low [acidity] at 0.01 [KOH] (whereas the type DF is a rather high at
0.15 and the bottle states that it may damage objectives in constant use -
it has lower auto-fluorescence than FF though). Cargille states that
Cedarwood oil has a KOH neutralisation number greater than these values (ie.
it is more acidic).

When used for objective immersion, the main things of interest in an oil
would be things like : refractive index, colour, fluorescent properties,
likelihood of damage to the lens cement, crystallisation when cold
(reversible), and whether the oil evaporates leaving a hard residue. The
metal outside parts of the objective generally have an anti-corrosion
finish, although here with our inverted microscopes spillage of corrosive
culture media has sometimes caused some damage at the base of the objective,
in particular corrosion of the area around the screw thread. The immersion
oils generally dissolve in things like ethyl alcohol. Rather like engine
oils for your car, one tends to stick with the manufacturers [typically
expensive] recommended one and not look too deeply into it's chemistry
(microscope manufacturer's will soak test their objectives immersed for 6
months or more in an immersion oil before recommending it).

Oil acids can attack the lens mounting cement [see Cargille's comments
below]. Plus the organic solvents used to remove lens immersion oil (like
Xylene and ethanol) have been implicated in lens cement damage - I use ether
generally for cleaning (and only clean rarely, when the lens is too dirty to
be gently cleaned by just a lens tissue). Modern [low acid content]
immersion oils supplied by the microscope manufacturer seem to be safe for
longer term contact with the objective and don't evaporate to a hard residue
(unlike say corn oil which leaves a very hard residue). Thus modern
microscope immersion oils are termed 'non-drying'.

Cargille quote immersion oil shelf lives of 10 years (FF) and 5 years (DF) -
and half that when opened.

On Cargille's site [http://www.cargille.com/immersionoilmicroscope.shtml]
John Cargille states:

"To the knowledge of the author, no immersion oil matches the optics of the
microscope perfectly. One of the closest matches is thickened cedarwood oil
which, for many years, was the most widely used, if not the only immersion
oil available. The disadvantageous properties of cedarwood oil are: high
absorption of blue and UV light, yellowing with age, a tendency to harden on
lenses due to uneven volatility, acidity, and changing viscosity (diluting
with solvent changes the index and dispersion). The synthetic immersion oils
eliminate many of the disadvantages cited above.

The acid value of immersion oil should be very low. The synthetics usually
have acid values lower than cedarwood oil.

High acidity can, in time, affect the condition of the metal parts of the
objective, and possibly more important, can cause deterioration of lens
cements. The lens cement is also a seal that prevents oil from penetrating
to the back of the lens. A crack or perforation in the cement draws oil by
capillary attraction and a thin film of oil slowly creeps over the back of
the objective lens; a poor image may develop without being immediately
noticed. When the microscopist realizes the image has deteriorated, he may
not realize the cause unless he has faced this problem before."

See Cargille's excellent site for more immersion oil info.
http://www.cargille.com/immersionoilmicroscope.shtml

Regards

Keith



----------------------------------------------------------
Dr Keith J Morris
Imaging Facilities Manager
Cell Biology Division
Institute of Ophthalmology
University College London
11-43 Bath Street
London EC1V 9EL

Tel: [phone]
Fax: [phone]
email: keith.morris-at-ucl.ac.uk

-----Original Message-----
X-from: ph2-at-sprynet.com [mailto:ph2-at-sprynet.com]
Sent: 22 June 2006 04:40
To: keith.morris-at-ucl.ac.uk

Regarding:

Name: raffaela

Education: Undergraduate College

Question: what is the pH of a cedarwood oilJuniperus virginiana) for oil
immersion objective? what is the pH of lemongrass oil(Cymbopogon citratus)?

1. There are three types of cedar oil:

Virginia cedarwood oil (Juniperus virginiana), Texas cedarwood oil
(Juniperus ashei or mexicana), and Western red cedar (Thuja plicata)

I have cedar oil (Texas) from Polysciences. Their MSDS does not list a pH.
Nor does Merck list one. The Sigma Aldrich MSDS does not list one, nor does
the Libety Nautral Products (Virginia), nor does JT Baker, nor does Well,
Naturally Products (Virginia & Texas & Western).

I will try to get a pH on my Texas cedar oil next week and report this at
least.

The constituents are mainly cedrene (a terpene) and cedral (cedar camphor).

2. regarding lemongrass oil:

I have found no pH. Well, Naturally Products does not have it in their
MSDS.

Constituents (as listed by Merck): 75-85% citral, methylheptenone,
citronellel, geraniol, limonene

3. I have a client that extracts, sells, distributes these oils and
will check for their data as well next week.

Tony

..........................................................................

90% of Risk Management is knowing where to place the decimal point...any
consultant can give you the other 10%â„



==============================Original Headers==============================
43, 27 -- From keith.morris-at-ucl.ac.uk Thu Jun 22 09:52:39 2006
43, 27 -- Received: from vscani-d.ucl.ac.uk (vscani-d.ucl.ac.uk
[144.82.108.132])
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-0500
==============================Original Headers==============================

[Boswell, Carl A - (cboswell)]

Regarding the aside about objectives being consumables:  I was informed by Zeiss when I told them that oil had leaked into our 63x Plan Apo that they consider objectives consumables, just like lens paper I guess.  It cost about 60% the cost of a new lens to have it reworked.  Check your service agreements/vendors and see what they say.  You might be surprised.

Carl

 

Carl A. Boswell, Ph.D.
Molecular and Cellular Biology
University of Arizona
520-954-7053
FAX [phone]

----- Original Message -----

From: [hidden email]

To: [hidden email]

Sent: Monday, March 23, 2009 3:18 AM

Subject: Re: Immersion Oil - 37oC

Hi all,

 

Using immersion oil at 37oC.

 

Zeiss sell a range of low fluorescence immersion oils: bottles of 20, 250 and 500 ml, unfortunately all filled with the same 518F oil. According to Zeiss it’s apparently ‘compatible’ with our Zeiss microscopes, but little extra information is offered on the Zeiss website.

 

The only info I can gleam from the web [other than the ubiquitous 518F safety data Sheet] is the following

Immersion oil "Immersol" 518 F fluorescence free, (ISO 8036-1/2, n(e)=1.518 (23°C), halogen-free)

 

So not much comment about 37oC use.  PeCon’s site [who supply most of Zeiss’s decent incubator stuff] have no opinion on the matter either.

 

The 518F oil safety sheet mentions “Use of the Substance / Preparation: For application information, consult the processing instructions. Processing instructions or technical

information sheet available on request.”. I’ll ask our knowledgeable local Zeiss Confocal rep for more details, as we are just installing a Zeiss/PeCon 37oC live cell incubator here on our Zeiss 510 MetaHead confocal.

Cargilee’s site in comparison is passionate about all things immersion oil:
http://www.cargille.com/immeroil.shtml
http://www.cargille.com/immeroilselection.shtml
and their excellent, if elderly, ‘primer’
http://www.cargille.com/immersionoilmicroscope.shtml

From postings to this server back in 2006, and Cargille’s site not mentioning ‘non-fluorescence’ for type 37, it appears that Cargille 37 probably autofluoresces, so it’s likely better for live cell phase-contrast [halogen lamp] transmission microscopy [but then often so are dry objectives]. I’ll stick with standard 518F immersion oil at 37oC though, it’s always imaged well enough on Zeiss microscopes at that temperature [and at least at 37oC it never became cloudy due to crystal precipitation]. Similarly the Cargille DF and FF seem fine to me at 37oC, and we stick ruthlessly with the same brand on each microscope. A lot of our live cell work is/was done with low power [20x Phase] air objectives though.

I do notice that immersion oil often ingresses into oil immersion objective internal optics with time, although strangely it rarely notices when imaging [like dirt on much of the internal optics, you must mostly focus through it]. One microscopy core manager even commented to me that he considers oil objectives ‘consumables’. The cost of ‘repair’ [often the same price as a new objective] is so high we frequently have to live with it [I advise users that our say 40x objective image quality is suspect for this reason]. In truth we don’t notice any drop in image quality when it must have happened, although you can see it ‘smeared’ on the optics when looking through the removed objective, using a magnifying glass, bright room light and a ‘cleaned’ objective top lens.

Old listerver posting on Cargille 37 attached below.

Regards

Keith

PS missed this link on the last post:

Dissolving crystals in Zeiss 518F oil
http://www.zeiss.com/C1256F8500454979/0/E8CD09DFA520787FC1256F86004B5FFB/$file/immersol-crystals.pdf

---------------------------------------------------------------------------
Dr Keith J. Morris,
Molecular Cytogenetics and Microscopy Core,
Laboratory 00/069 and 00/070,
The Wellcome Trust Centre for Human Genetics,
Roosevelt Drive,
Oxford  OX3 7BN,
United Kingdom.

Telephone:  [phone]
Email:  [hidden email]
Web-pages: http://www.well.ox.ac.uk/cytogenetics/

 

Subject:	Re: Fluorescence from Type 37 Immersion Oil
From:	Vitaly Boyko <[hidden email]>
Reply-To:	Vitaly Boyko <[hidden email]>
Date:	Wed, 16 Aug 2006 10:23:20 -0400
Content-Type:	text/plain

 

Search the CONFOCAL archive at

http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

PS What would be more appropriate to compare is Zeiss 518F (not 518C!) with 

the Nikon type A or Cargille type DF.

Hi Jason,

I wonder why you have tested the Cargille FF imm. oil, as the refractive

index of FF oil is too far off for the standard glass (1.518) ?????!!!!!

Actually, I haven't noticed the "very bad" autofluorescence of the Cargille

type DF versus Zeiss 518 in the CFP, YFP or FRET channels. The difference is

less than 10%.

The problem with the Nikon oil is its dispenser and/or viscosity - it is

often annoying "fighting" with the air bubbles.

What is about the Invitrogen/Mol.Probes mounting media autofluorescence?

I am fed up with the irreproducibility of the background autofluorescence

using the ProlongGold mounting media!!! That is a serious problem compared

to the issue of the immersion oils.

Cheers,

Vitaly

NCI-Frederick

----- Original Message ----- 

From: "Kilgore, Jason" <[hidden email]>

To: <[hidden email]>

Sent: Tuesday, August 15, 2006 7:26 PM

Subject: Re: Fluorescence from Type 37 Immersion Oil

> Search the CONFOCAL archive at

> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

> 

> Back in 2000 I performed a comparison of four different immersion oils

> (Zeiss 518C, Nikon Type A, Cargille Type DF, and Cargille Type FF) for

> autofluorescence (using DAPI, FITC, acridine orange, and TRITC filter

> sets) and general resolution at 60x on a Nikon E400.  I didn't test

> Cargille 37.

> 

> Green autofluorescence tended to be worst for all four, but was least

> for Cargille FF, followed by Nikon Type A, then Zeiss, then (very bad)

> Cargille DF.  However, the resolution was unacceptable for Cargille FF.

> Autofluorescence in the other channels was moderate (DAPI, TRITC) to

> very low (acridine orange), with the same comparative trend applying.

> 

> Thus, ever since we have been using Nikon Type A with no complaints for

> most of our imaging.

> 

> Feel free to contact me personally and I can send you the data.

> 

> Jason

> 

> Jason A. Kilgore

> Cell Biology / Histology

> Molecular Probes/Invitrogen

> [hidden email]

> 

> 

> -----Original Message-----

> From: Confocal Microscopy List [mailto:[hidden email]] On

> Behalf Of S. Brunet

> Sent: Tuesday, August 15, 2006 11:19 AM

> To: [hidden email]

> Subject: Fluorescence from Type 37 Immersion Oil

> 

> Search the CONFOCAL archive at

> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

> 

> Hello to all,

> 

> We recently began using the Cargille 37 oil with the temperature chamber

> on the

> microscope.  I found significant fluorescence from the oil when exposed

> to the

> HBO lamp (I could not see the eYFP because of the blue background).

> 

> So I used the 20x air objective and I was able to obtain an image from

> the

> fluorescence from a drop of 37 oil placed on a coverslip as a sample

> (excitation at 488 or 458nm and pinhole at the usual confocal setting).

> 

> Is this the norm?

> 

> Regards,

> Sophie

> _______________________________________

> Sophie M.K. Brunet, Ph.D.

> Research Officer

> Optical Spectroscopist, Laser Systems and Applications

> [hidden email]

> 306-966-1719 (office)   306-966-1702 (fax)

> _______________________________________

> Saskatchewan Structural Sciences Centre

> University of Saskatchewan

> Thorvaldson Bldg.

> 110 Science Place

> Saskatoon, SK   S7N 5C9

> 

 

[Chris Wood-5]

Won't this hysteresis play havoc with software-based autofocus routines?

Any ideas on compatibility with such techniques?

Saludos

Chris

IBt/UNAM

On Mon, 23 Mar 2009 08:56:00 -0400, John Oreopoulos
<[hidden email]> wrote:

>We purchased a bottle of this product a few months back and I've used  
>it a few times when doing live-cell imaging. One thing I noticed that  
>was different when using this compared to regular immersion oil for  
>fluorescence was that there was a short lag time in terms of  
>adjusting the focus of the microscope and the image coming into focus  
>on my preview computer screen (and in the eyepieces). I guess this  
>has to do with the viscosity of the oil. It took a while to get  
>adjusted to this because I would keep overshooting the fine focus,  
>but other than that it works great (no commercial interest).
>
>John Oreopoulos
>
>
>On 23-Mar-09, at 6:42 AM, Christophe Leterrier wrote:
>
>> I would like to point out that Cargille has a new oil for fluorescence
>> imaging at 37°C, called 37DF, that has very low auto fluorescence. It
>> is not on their website, but I could get the product pdf sent to me by
>> a Cargille person, so I can send it if someone is interested. I'm
>> trying to get some here in France, but none of the french Cargille
>> distributors has heard about it yet... It has been found to be quite
>> good for fluorescence imaging of live cells at 37°C, according to this
>> paper in J Microscopy :
>>
>> Immersion oil for high-resolution live-cell imaging at 37 degrees C:
>> optical and physical characteristics.
>> Oomen LC, Sacher R, Brocks HH, Zwier JM, Brakenhoff GJ, Jalink K.
>> J Microsc. 2008 Nov;232(2):353-61.
>>
>>
>> On Mon, Mar 23, 2009 at 11:18, Keith Morris  
>> <[hidden email]> wrote:
>>>
>>> Hi all,
>>>
>>>
>>>
>>> Using immersion oil at 37oC.
>>>
>>>
>>>
>>> Zeiss sell a range of low fluorescence immersion oils: bottles of  
>>> 20, 250 and 500 ml, unfortunately all filled with the same 518F  
>>> oil. According to Zeiss it’s apparently ‘compatible’ with our  
>>> Zeiss microscopes, but little extra information is offered on the  
>>> Zeiss website.
>>>
>>>
>>>
>>> The only info I can gleam from the web [other than the ubiquitous  
>>> 518F safety data Sheet] is the following
>>>
>>> Immersion oil "Immersol" 518 F fluorescence free, (ISO 8036-1/2, n
>>> (e)=1.518 (23°C), halogen-free)
>>>
>>>
>>>
>>> So not much comment about 37oC use.  PeCon’s site [who supply most  
>>> of Zeiss’s decent incubator stuff] have no opinion on the matter  
>>> either.
>>>
>>>
>>>
>>> The 518F oil safety sheet mentions “Use of the Substance /  
>>> Preparation: For application information, consult the processing  
>>> instructions. Processing instructions or technical
>>>
>>> information sheet available on request.”. I’ll ask our  
>>> knowledgeable local Zeiss Confocal rep for more details, as we are  
>>> just installing a Zeiss/PeCon 37oC live cell incubator here on our  
>>> Zeiss 510 MetaHead confocal.
>>>
>>> Cargilee’s site in comparison is passionate about all things  
>>> immersion oil:
>>> http://www.cargille.com/immeroil.shtml
>>> http://www.cargille.com/immeroilselection.shtml
>>> and their excellent, if elderly, ‘primer’
>>> http://www.cargille.com/immersionoilmicroscope.shtml
>>>
>>> From postings to this server back in 2006, and Cargille’s site not  
>>> mentioning ‘non-fluorescence’ for type 37, it appears that  
>>> Cargille 37 probably autofluoresces, so it’s likely better for  
>>> live cell phase-contrast [halogen lamp] transmission microscopy  
>>> [but then often so are dry objectives]. I’ll stick with standard  
>>> 518F immersion oil at 37oC though, it’s always imaged well enough  
>>> on Zeiss microscopes at that temperature [and at least at 37oC it  
>>> never became cloudy due to crystal precipitation]. Similarly the  
>>> Cargille DF and FF seem fine to me at 37oC, and we stick  
>>> ruthlessly with the same brand on each microscope. A lot of our  
>>> live cell work is/was done with low power [20x Phase] air  
>>> objectives though.
>>>
>>> I do notice that immersion oil often ingresses into oil immersion  
>>> objective internal optics with time, although strangely it rarely  
>>> notices when imaging [like dirt on much of the internal optics,  
>>> you must mostly focus through it]. One microscopy core manager  
>>> even commented to me that he considers oil objectives  
>>> ‘consumables’. The cost of ‘repair’ [often the same price as a new  
>>> objective] is so high we frequently have to live with it [I advise  
>>> users that our say 40x objective image quality is suspect for this  
>>> reason]. In truth we don’t notice any drop in image quality when  
>>> it must have happened, although you can see it ‘smeared’ on the  
>>> optics when looking through the removed objective, using a  
>>> magnifying glass, bright room light and a ‘cleaned’ objective top  
>>> lens.
>>>
>>> Old listerver posting on Cargille 37 attached below.
>>>
>>> Regards
>>>
>>> Keith
>>>
>>> PS missed this link on the last post:
>>>
>>> Dissolving crystals in Zeiss 518F oil
>>> http://www.zeiss.com/C1256F8500454979/0/ 
>>> E8CD09DFA520787FC1256F86004B5FFB/$file/immersol-crystals.pdf
>>>
>>> ---------------------------------------------------------------------
>>> ------
>>> Dr Keith J. Morris,
>>> Molecular Cytogenetics and Microscopy Core,
>>> Laboratory 00/069 and 00/070,
>>> The Wellcome Trust Centre for Human Genetics,
>>> Roosevelt Drive,
>>> Oxford  OX3 7BN,
>>> United Kingdom.
>>>
>>> Telephone:  [phone]
>>> Email:  [hidden email]
>>> Web-pages: http://www.well.ox.ac.uk/cytogenetics/
>>>
>>>
>>>
>>> Subject:
>>>
>>> Re: Fluorescence from Type 37 Immersion Oil
>>>
>>> From:
>>>
>>> Vitaly Boyko <[hidden email]>
>>>
>>> Reply-To:
>>>
>>> Vitaly Boyko <[hidden email]>
>>>
>>> Date:
>>>
>>> Wed, 16 Aug 2006 10:23:20 -0400
>>>
>>> Content-Type:
>>>
>>> text/plain
>>>
>>>
>>>
>>> Search the CONFOCAL archive at
>>>
>>> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>>>
>>>
>>>
>>> PS What would be more appropriate to compare is Zeiss 518F (not  
>>> 518C!) with
>>>
>>> the Nikon type A or Cargille type DF.
>>>
>>>
>>>
>>>
>>>
>>>
>>>
>>> Hi Jason,
>>>
>>>
>>>
>>> I wonder why you have tested the Cargille FF imm. oil, as the  
>>> refractive
>>>
>>> index of FF oil is too far off for the standard glass  
>>> (1.518) ?????!!!!!
>>>
>>>
>>>
>>> Actually, I haven't noticed the "very bad" autofluorescence of the  
>>> Cargille
>>>
>>> type DF versus Zeiss 518 in the CFP, YFP or FRET channels. The  
>>> difference is
>>>
>>> less than 10%.
>>>
>>>
>>>
>>> The problem with the Nikon oil is its dispenser and/or viscosity -  
>>> it is
>>>
>>> often annoying "fighting" with the air bubbles.
>>>
>>>
>>>
>>> What is about the Invitrogen/Mol.Probes mounting media  
>>> autofluorescence?
>>>
>>>
>>>
>>> I am fed up with the irreproducibility of the background  
>>> autofluorescence
>>>
>>> using the ProlongGold mounting media!!! That is a serious problem  
>>> compared
>>>
>>> to the issue of the immersion oils.
>>>
>>>
>>>
>>> Cheers,
>>>
>>>
>>>
>>> Vitaly
>>>
>>>
>>>
>>> NCI-Frederick
>>>
>>>
>>>
>>>
>>>
>>> ----- Original Message -----
>>>
>>> From: "Kilgore, Jason" <[hidden email]>
>>>
>>> To: <[hidden email]>
>>>
>>> Sent: Tuesday, August 15, 2006 7:26 PM
>>>
>>> Subject: Re: Fluorescence from Type 37 Immersion Oil
>>>
>>>
>>>
>>>
>>>
>>>> Search the CONFOCAL archive at
>>>
>>>> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>>>
>>>>
>>>
>>>> Back in 2000 I performed a comparison of four different immersion  
>>>> oils
>>>
>>>> (Zeiss 518C, Nikon Type A, Cargille Type DF, and Cargille Type  
>>>> FF) for
>>>
>>>> autofluorescence (using DAPI, FITC, acridine orange, and TRITC  
>>>> filter
>>>
>>>> sets) and general resolution at 60x on a Nikon E400.  I didn't test
>>>
>>>> Cargille 37.
>>>
>>>>
>>>
>>>> Green autofluorescence tended to be worst for all four, but was  
>>>> least
>>>
>>>> for Cargille FF, followed by Nikon Type A, then Zeiss, then (very  
>>>> bad)
>>>
>>>> Cargille DF.  However, the resolution was unacceptable for  
>>>> Cargille FF.
>>>
>>>> Autofluorescence in the other channels was moderate (DAPI, TRITC) to
>>>
>>>> very low (acridine orange), with the same comparative trend  
>>>> applying.
>>>
>>>>
>>>
>>>> Thus, ever since we have been using Nikon Type A with no  
>>>> complaints for
>>>
>>>> most of our imaging.
>>>
>>>>
>>>
>>>> Feel free to contact me personally and I can send you the data.
>>>
>>>>
>>>
>>>> Jason
>>>
>>>>
>>>
>>>> Jason A. Kilgore
>>>
>>>> Cell Biology / Histology
>>>
>>>> Molecular Probes/Invitrogen
>>>
>>>> [hidden email]
>>>
>>>>
>>>
>>>>
>>>
>>>> -----Original Message-----
>>>
>>>> From: Confocal Microscopy List  
>>>> [mailto:[hidden email]] On
>>>
>>>> Behalf Of S. Brunet
>>>
>>>> Sent: Tuesday, August 15, 2006 11:19 AM
>>>
>>>> To: [hidden email]
>>>
>>>> Subject: Fluorescence from Type 37 Immersion Oil
>>>
>>>>
>>>
>>>> Search the CONFOCAL archive at
>>>
>>>> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>>>
>>>>
>>>
>>>> Hello to all,
>>>
>>>>
>>>
>>>> We recently began using the Cargille 37 oil with the temperature  
>>>> chamber
>>>
>>>> on the
>>>
>>>> microscope.  I found significant fluorescence from the oil when  
>>>> exposed
>>>
>>>> to the
>>>
>>>> HBO lamp (I could not see the eYFP because of the blue background).
>>>
>>>>
>>>
>>>> So I used the 20x air objective and I was able to obtain an image  
>>>> from
>>>
>>>> the
>>>
>>>> fluorescence from a drop of 37 oil placed on a coverslip as a sample
>>>
>>>> (excitation at 488 or 458nm and pinhole at the usual confocal  
>>>> setting).
>>>
>>>>
>>>
>>>> Is this the norm?
>>>
>>>>
>>>
>>>> Regards,
>>>
>>>> Sophie
>>>
>>>> _______________________________________
>>>
>>>> Sophie M.K. Brunet, Ph.D.
>>>
>>>> Research Officer
>>>
>>>> Optical Spectroscopist, Laser Systems and Applications
>>>
>>>> [hidden email]
>>>
>>>> 306-966-1719 (office)   306-966-1702 (fax)
>>>
>>>> _______________________________________
>>>
>>>> Saskatchewan Structural Sciences Centre
>>>
>>>> University of Saskatchewan
>>>
>>>> Thorvaldson Bldg.
>>>
>>>> 110 Science Place
>>>
>>>> Saskatoon, SK   S7N 5C9
>>>
>>>>
>>>
>>>

[Larson Jeffrey M.]

Dear list,

This is a commercial response.

Nikon's 'Perfect Focus System' corrects for focus drift by moving an internal lens located inside the PFS unit.  Because the nosepiece does not move, viscosity of the immersion medium does not present a problem.  Regarding Nikon immersion oil, my experience is that bubbles usually disappear after a few seconds and stubborn bubbles can be burst by touching them with the tip the applicator.  Bubbles are rarely seen when the 8cc applicator is used.

Jeffrey Larson
Product Manager
Nikon Confocal Systems
Nikon Instruments Inc.
[phone]

-----Original Message-----
From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Chris Wood
Sent: Tuesday, March 24, 2009 11:39 AM
To: [hidden email]
Subject: Re: Immersion Oil - 37oC

Won't this hysteresis play havoc with software-based autofocus routines?

Any ideas on compatibility with such techniques?

Saludos

Chris

IBt/UNAM

On Mon, 23 Mar 2009 08:56:00 -0400, John Oreopoulos
<[hidden email]> wrote:

>We purchased a bottle of this product a few months back and I've used  
>it a few times when doing live-cell imaging. One thing I noticed that  
>was different when using this compared to regular immersion oil for  
>fluorescence was that there was a short lag time in terms of  
>adjusting the focus of the microscope and the image coming into focus  
>on my preview computer screen (and in the eyepieces). I guess this  
>has to do with the viscosity of the oil. It took a while to get  
>adjusted to this because I would keep overshooting the fine focus,  
>but other than that it works great (no commercial interest).
>
>John Oreopoulos
>
>
>On 23-Mar-09, at 6:42 AM, Christophe Leterrier wrote:
>
>> I would like to point out that Cargille has a new oil for fluorescence
>> imaging at 37°C, called 37DF, that has very low auto fluorescence. It
>> is not on their website, but I could get the product pdf sent to me by
>> a Cargille person, so I can send it if someone is interested. I'm
>> trying to get some here in France, but none of the french Cargille
>> distributors has heard about it yet... It has been found to be quite
>> good for fluorescence imaging of live cells at 37°C, according to this
>> paper in J Microscopy :
>>
>> Immersion oil for high-resolution live-cell imaging at 37 degrees C:
>> optical and physical characteristics.
>> Oomen LC, Sacher R, Brocks HH, Zwier JM, Brakenhoff GJ, Jalink K.
>> J Microsc. 2008 Nov;232(2):353-61.
>>
>>
>> On Mon, Mar 23, 2009 at 11:18, Keith Morris  
>> <[hidden email]> wrote:
>>>
>>> Hi all,
>>>
>>>
>>>
>>> Using immersion oil at 37oC.
>>>
>>>
>>>
>>> Zeiss sell a range of low fluorescence immersion oils: bottles of  
>>> 20, 250 and 500 ml, unfortunately all filled with the same 518F  
>>> oil. According to Zeiss it's apparently 'compatible' with our  
>>> Zeiss microscopes, but little extra information is offered on the  
>>> Zeiss website.
>>>
>>>
>>>
>>> The only info I can gleam from the web [other than the ubiquitous  
>>> 518F safety data Sheet] is the following
>>>
>>> Immersion oil "Immersol" 518 F fluorescence free, (ISO 8036-1/2, n
>>> (e)=1.518 (23°C), halogen-free)
>>>
>>>
>>>
>>> So not much comment about 37oC use.  PeCon's site [who supply most  
>>> of Zeiss's decent incubator stuff] have no opinion on the matter  
>>> either.
>>>
>>>
>>>
>>> The 518F oil safety sheet mentions "Use of the Substance /  
>>> Preparation: For application information, consult the processing  
>>> instructions. Processing instructions or technical
>>>
>>> information sheet available on request.". I'll ask our  
>>> knowledgeable local Zeiss Confocal rep for more details, as we are  
>>> just installing a Zeiss/PeCon 37oC live cell incubator here on our  
>>> Zeiss 510 MetaHead confocal.
>>>
>>> Cargilee's site in comparison is passionate about all things  
>>> immersion oil:
>>> http://www.cargille.com/immeroil.shtml
>>> http://www.cargille.com/immeroilselection.shtml
>>> and their excellent, if elderly, 'primer'
>>> http://www.cargille.com/immersionoilmicroscope.shtml
>>>
>>> From postings to this server back in 2006, and Cargille's site not  
>>> mentioning 'non-fluorescence' for type 37, it appears that  
>>> Cargille 37 probably autofluoresces, so it's likely better for  
>>> live cell phase-contrast [halogen lamp] transmission microscopy  
>>> [but then often so are dry objectives]. I'll stick with standard  
>>> 518F immersion oil at 37oC though, it's always imaged well enough  
>>> on Zeiss microscopes at that temperature [and at least at 37oC it  
>>> never became cloudy due to crystal precipitation]. Similarly the  
>>> Cargille DF and FF seem fine to me at 37oC, and we stick  
>>> ruthlessly with the same brand on each microscope. A lot of our  
>>> live cell work is/was done with low power [20x Phase] air  
>>> objectives though.
>>>
>>> I do notice that immersion oil often ingresses into oil immersion  
>>> objective internal optics with time, although strangely it rarely  
>>> notices when imaging [like dirt on much of the internal optics,  
>>> you must mostly focus through it]. One microscopy core manager  
>>> even commented to me that he considers oil objectives  
>>> 'consumables'. The cost of 'repair' [often the same price as a new  
>>> objective] is so high we frequently have to live with it [I advise  
>>> users that our say 40x objective image quality is suspect for this  
>>> reason]. In truth we don't notice any drop in image quality when  
>>> it must have happened, although you can see it 'smeared' on the  
>>> optics when looking through the removed objective, using a  
>>> magnifying glass, bright room light and a 'cleaned' objective top  
>>> lens.
>>>
>>> Old listerver posting on Cargille 37 attached below.
>>>
>>> Regards
>>>
>>> Keith
>>>
>>> PS missed this link on the last post:
>>>
>>> Dissolving crystals in Zeiss 518F oil
>>> http://www.zeiss.com/C1256F8500454979/0/ 
>>> E8CD09DFA520787FC1256F86004B5FFB/$file/immersol-crystals.pdf
>>>
>>> ---------------------------------------------------------------------
>>> ------
>>> Dr Keith J. Morris,
>>> Molecular Cytogenetics and Microscopy Core,
>>> Laboratory 00/069 and 00/070,
>>> The Wellcome Trust Centre for Human Genetics,
>>> Roosevelt Drive,
>>> Oxford  OX3 7BN,
>>> United Kingdom.
>>>
>>> Telephone:  [phone]
>>> Email:  [hidden email]
>>> Web-pages: http://www.well.ox.ac.uk/cytogenetics/
>>>
>>>
>>>
>>> Subject:
>>>
>>> Re: Fluorescence from Type 37 Immersion Oil
>>>
>>> From:
>>>
>>> Vitaly Boyko <[hidden email]>
>>>
>>> Reply-To:
>>>
>>> Vitaly Boyko <[hidden email]>
>>>
>>> Date:
>>>
>>> Wed, 16 Aug 2006 10:23:20 -0400
>>>
>>> Content-Type:
>>>
>>> text/plain
>>>
>>>
>>>
>>> Search the CONFOCAL archive at
>>>
>>> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>>>
>>>
>>>
>>> PS What would be more appropriate to compare is Zeiss 518F (not  
>>> 518C!) with
>>>
>>> the Nikon type A or Cargille type DF.
>>>
>>>
>>>
>>>
>>>
>>>
>>>
>>> Hi Jason,
>>>
>>>
>>>
>>> I wonder why you have tested the Cargille FF imm. oil, as the  
>>> refractive
>>>
>>> index of FF oil is too far off for the standard glass  
>>> (1.518) ?????!!!!!
>>>
>>>
>>>
>>> Actually, I haven't noticed the "very bad" autofluorescence of the  
>>> Cargille
>>>
>>> type DF versus Zeiss 518 in the CFP, YFP or FRET channels. The  
>>> difference is
>>>
>>> less than 10%.
>>>
>>>
>>>
>>> The problem with the Nikon oil is its dispenser and/or viscosity -  
>>> it is
>>>
>>> often annoying "fighting" with the air bubbles.
>>>
>>>
>>>
>>> What is about the Invitrogen/Mol.Probes mounting media  
>>> autofluorescence?
>>>
>>>
>>>
>>> I am fed up with the irreproducibility of the background  
>>> autofluorescence
>>>
>>> using the ProlongGold mounting media!!! That is a serious problem  
>>> compared
>>>
>>> to the issue of the immersion oils.
>>>
>>>
>>>
>>> Cheers,
>>>
>>>
>>>
>>> Vitaly
>>>
>>>
>>>
>>> NCI-Frederick
>>>
>>>
>>>
>>>
>>>
>>> ----- Original Message -----
>>>
>>> From: "Kilgore, Jason" <[hidden email]>
>>>
>>> To: <[hidden email]>
>>>
>>> Sent: Tuesday, August 15, 2006 7:26 PM
>>>
>>> Subject: Re: Fluorescence from Type 37 Immersion Oil
>>>
>>>
>>>
>>>
>>>
>>>> Search the CONFOCAL archive at
>>>
>>>> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>>>
>>>>
>>>
>>>> Back in 2000 I performed a comparison of four different immersion  
>>>> oils
>>>
>>>> (Zeiss 518C, Nikon Type A, Cargille Type DF, and Cargille Type  
>>>> FF) for
>>>
>>>> autofluorescence (using DAPI, FITC, acridine orange, and TRITC  
>>>> filter
>>>
>>>> sets) and general resolution at 60x on a Nikon E400.  I didn't test
>>>
>>>> Cargille 37.
>>>
>>>>
>>>
>>>> Green autofluorescence tended to be worst for all four, but was  
>>>> least
>>>
>>>> for Cargille FF, followed by Nikon Type A, then Zeiss, then (very  
>>>> bad)
>>>
>>>> Cargille DF.  However, the resolution was unacceptable for  
>>>> Cargille FF.
>>>
>>>> Autofluorescence in the other channels was moderate (DAPI, TRITC) to
>>>
>>>> very low (acridine orange), with the same comparative trend  
>>>> applying.
>>>
>>>>
>>>
>>>> Thus, ever since we have been using Nikon Type A with no  
>>>> complaints for
>>>
>>>> most of our imaging.
>>>
>>>>
>>>
>>>> Feel free to contact me personally and I can send you the data.
>>>
>>>>
>>>
>>>> Jason
>>>
>>>>
>>>
>>>> Jason A. Kilgore
>>>
>>>> Cell Biology / Histology
>>>
>>>> Molecular Probes/Invitrogen
>>>
>>>> [hidden email]
>>>
>>>>
>>>
>>>>
>>>
>>>> -----Original Message-----
>>>
>>>> From: Confocal Microscopy List  
>>>> [mailto:[hidden email]] On
>>>
>>>> Behalf Of S. Brunet
>>>
>>>> Sent: Tuesday, August 15, 2006 11:19 AM
>>>
>>>> To: [hidden email]
>>>
>>>> Subject: Fluorescence from Type 37 Immersion Oil
>>>
>>>>
>>>
>>>> Search the CONFOCAL archive at
>>>
>>>> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>>>
>>>>
>>>
>>>> Hello to all,
>>>
>>>>
>>>
>>>> We recently began using the Cargille 37 oil with the temperature  
>>>> chamber
>>>
>>>> on the
>>>
>>>> microscope.  I found significant fluorescence from the oil when  
>>>> exposed
>>>
>>>> to the
>>>
>>>> HBO lamp (I could not see the eYFP because of the blue background).
>>>
>>>>
>>>
>>>> So I used the 20x air objective and I was able to obtain an image  
>>>> from
>>>
>>>> the
>>>
>>>> fluorescence from a drop of 37 oil placed on a coverslip as a sample
>>>
>>>> (excitation at 488 or 458nm and pinhole at the usual confocal  
>>>> setting).
>>>
>>>>
>>>
>>>> Is this the norm?
>>>
>>>>
>>>
>>>> Regards,
>>>
>>>> Sophie
>>>
>>>> _______________________________________
>>>
>>>> Sophie M.K. Brunet, Ph.D.
>>>
>>>> Research Officer
>>>
>>>> Optical Spectroscopist, Laser Systems and Applications
>>>
>>>> [hidden email]
>>>
>>>> 306-966-1719 (office)   306-966-1702 (fax)
>>>
>>>> _______________________________________
>>>
>>>> Saskatchewan Structural Sciences Centre
>>>
>>>> University of Saskatchewan
>>>
>>>> Thorvaldson Bldg.
>>>
>>>> 110 Science Place
>>>
>>>> Saskatoon, SK   S7N 5C9
>>>
>>>>
>>>
>>>

[James Pawley]

Hi all,

In Chapter 11 of the Handbook, Rimas Juskaitis goes into some detail about measurements that he made on the relatively large changes in RI that can occur when immersion oils are use at other than room temp.

The bottom line is that it can be very finicky: even one deg C can make a noticeable difference. In addition,  once the oil has the WRONG RI, then the amount of spherical aberration produced will depend on the free working distance (i.e., it may look better or worse depending on the depth that you are focused into the specimen).

The solution as the Agard/Sedat group found years ago is to always include sub-resolution beads in your specimen and focus up-and-down on these in widefield to make sure that the point image becomes blurry in the same way, whether you focus up or down from best focus (i.e., blurry rings in both direction, not rings in one direction and blobs in the other).

Agard and Sedat were early users of Cargille oils mixed to the specific RI to give best results under the actual imaging conditions.

If trying to do the same with a water objective looking into a specimen with a higher RI than that of water, you can adjust the RI of the coupling liquid to be the same as that of the specimen with mixtures of water and glycerol, but then you will have to use the correction collar (and coverslip thickness) to compensate for the fact that you are no longer just looking into water.

Cheers,

Jim P.

              **********************************************
Prof. James B. Pawley,                                          Ph.  608-263-3147 
Room 223, Zoology Research Building,                                  FAX  [phone]
1117 Johnson Ave., Madison, WI, 53706                                [hidden email]
3D Microscopy of Living Cells Course, June 13-25, 2009, UBC, Vancouver Canada

Info: http://www.3dcourse.ubc.ca/                Applications still being accepted

               "If it ain't diffraction, it must be statistics." Anon.

Hi all,

 

Using immersion oil at 37oC.

 

Zeiss sell a range of low fluorescence immersion oils: bottles of 20, 250 and 500 ml, unfortunately all filled with the same 518F oil. According to Zeiss it's apparently 'compatible' with our Zeiss microscopes, but little extra information is offered on the Zeiss website.

 

The only info I can gleam from the web [other than the ubiquitous 518F safety data Sheet] is the following

Immersion oil "Immersol" 518 F fluorescence free, (ISO 8036-1/2, n(e)=1.518 (23°C), halogen-free)

 

So not much comment about 37oC use.  PeCon's site [who supply most of Zeiss's decent incubator stuff] have no opinion on the matter either.

 

The 518F oil safety sheet mentions "Use of the Substance / Preparation: For application information, consult the processing instructions. Processing instructions or technical

information sheet available on request.". I'll ask our knowledgeable local Zeiss Confocal rep for more details, as we are just installing a Zeiss/PeCon 37oC live cell incubator here on our Zeiss 510 MetaHead confocal.

Cargilee's site in comparison is passionate about all things immersion oil:

http://www.cargille.com/immeroil.shtml
http://www.cargille.com/immeroilselection.shtml
and their excellent, if elderly, 'primer'
http://www.cargille.com/immersionoilmicroscope.shtml

From postings to this server back in 2006, and Cargille's site not mentioning 'non-fluorescence' for type 37, it appears that Cargille 37 probably autofluoresces, so it's likely better for live cell phase-contrast [halogen lamp] transmission microscopy [but then often so are dry objectives]. I'll stick with standard 518F immersion oil at 37oC though, it's always imaged well enough on Zeiss microscopes at that temperature [and at least at 37oC it never became cloudy due to crystal precipitation]. Similarly the Cargille DF and FF seem fine to me at 37oC, and we stick ruthlessly with the same brand on each microscope. A lot of our live cell work is/was done with low power [20x Phase] air objectives though.

I do notice that immersion oil often ingresses into oil immersion objective internal optics with time, although strangely it rarely notices when imaging [like dirt on much of the internal optics, you must mostly focus through it]. One microscopy core manager even commented to me that he considers oil objectives 'consumables'. The cost of 'repair' [often the same price as a new objective] is so high we frequently have to live with it [I advise users that our say 40x objective image quality is suspect for this reason]. In truth we don't notice any drop in image quality when it must have happened, although you can see it 'smeared' on the optics when looking through the removed objective, using a magnifying glass, bright room light and a 'cleaned' objective top lens.

Old listerver posting on Cargille 37 attached below.

Regards

Keith

PS missed this link on the last post:

Dissolving crystals in Zeiss 518F oil
http://www.zeiss.com/C1256F8500454979/0/E8CD09DFA520787FC1256F86004B5FFB/$file/immersol-crystals.pdf

---------------------------------------------------------------------------
Dr Keith J. Morris,
Molecular Cytogenetics and Microscopy Core,
Laboratory 00/069 and 00/070,
The Wellcome Trust Centre for Human Genetics,
Roosevelt Drive,
Oxford  OX3 7BN,
United Kingdom.

Telephone:  [phone]
Email:  [hidden email]
Web-pages: http://www.well.ox.ac.uk/cytogenetics/

 

Subject:

Re: Fluorescence from Type 37 Immersion Oil

From:

Vitaly Boyko <[hidden email]>

Reply-To:

Vitaly Boyko <[hidden email]>

Date:

Wed, 16 Aug 2006 10:23:20 -0400

Content-Type:

text/plain

 

Search the CONFOCAL archive at

http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

 

PS What would be more appropriate to compare is Zeiss 518F (not 518C!) with

the Nikon type A or Cargille type DF.

 

 

 

Hi Jason,

 

I wonder why you have tested the Cargille FF imm. oil, as the refractive

index of FF oil is too far off for the standard glass (1.518) ?????!!!!!

 

Actually, I haven't noticed the "very bad" autofluorescence of the Cargille

type DF versus Zeiss 518 in the CFP, YFP or FRET channels. The difference is

less than 10%.

 

The problem with the Nikon oil is its dispenser and/or viscosity - it is

often annoying "fighting" with the air bubbles.

 

What is about the Invitrogen/Mol.Probes mounting media autofluorescence?

 

I am fed up with the irreproducibility of the background autofluorescence

using the ProlongGold mounting media!!! That is a serious problem compared

to the issue of the immersion oils.

 

Cheers,

 

Vitaly

 

NCI-Frederick

 

 

----- Original Message -----

From: "Kilgore, Jason" <[hidden email]>

To: <[hidden email]>

Sent: Tuesday, August 15, 2006 7:26 PM

Subject: Re: Fluorescence from Type 37 Immersion Oil

 

 

> Search the CONFOCAL archive at

> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

> 

> Back in 2000 I performed a comparison of four different immersion oils

> (Zeiss 518C, Nikon Type A, Cargille Type DF, and Cargille Type FF) for

> autofluorescence (using DAPI, FITC, acridine orange, and TRITC filter

> sets) and general resolution at 60x on a Nikon E400.  I didn't test

> Cargille 37.

> 

> Green autofluorescence tended to be worst for all four, but was least

> for Cargille FF, followed by Nikon Type A, then Zeiss, then (very bad)

> Cargille DF.  However, the resolution was unacceptable for Cargille FF.

> Autofluorescence in the other channels was moderate (DAPI, TRITC) to

> very low (acridine orange), with the same comparative trend applying.

> 

> Thus, ever since we have been using Nikon Type A with no complaints for

> most of our imaging.

> 

> Feel free to contact me personally and I can send you the data.

> 

> Jason

> 

> Jason A. Kilgore

> Cell Biology / Histology

> Molecular Probes/Invitrogen

> [hidden email]

> 

> 

> -----Original Message-----

> From: Confocal Microscopy List [mailto:[hidden email]] On

> Behalf Of S. Brunet

> Sent: Tuesday, August 15, 2006 11:19 AM

> To: [hidden email]

> Subject: Fluorescence from Type 37 Immersion Oil

> 

> Search the CONFOCAL archive at

> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

> 

> Hello to all,

> 

> We recently began using the Cargille 37 oil with the temperature chamber

> on the

> microscope.  I found significant fluorescence from the oil when exposed

> to the

> HBO lamp (I could not see the eYFP because of the blue background).

> 

> So I used the 20x air objective and I was able to obtain an image from

> the

> fluorescence from a drop of 37 oil placed on a coverslip as a sample

> (excitation at 488 or 458nm and pinhole at the usual confocal setting).

> 

> Is this the norm?

> 

> Regards,

> Sophie

> _______________________________________

> Sophie M.K. Brunet, Ph.D.

> Research Officer

> Optical Spectroscopist, Laser Systems and Applications

> [hidden email]

> 306-966-1719 (office)   306-966-1702 (fax)

> _______________________________________

> Saskatchewan Structural Sciences Centre

> University of Saskatchewan

> Thorvaldson Bldg.

> 110 Science Place

> Saskatoon, SK   S7N 5C9

>

 

--

[Jeremy Adler-2]

Temperature and water objectives.

 

It seems that the RI of water changes little over the range RT-37C

 

But does anyone have figures for the water replacement fluid provided by Zeiss.

It is specified at 1.333 at 23C.

 

 

 

Dr Jeremy Adler

F451a

Cell Biologi

Wenner-Gren Inst.

The Arhenius Lab

Stockholm University

S-106 91 Stockholm

Sweden

tel [phone]

---

From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of James Pawley
Sent: den 24 mars 2009 23:32
To: [hidden email]
Subject: Re: Immersion Oil - 37oC

 

Hi all,

 

In Chapter 11 of the Handbook, Rimas Juskaitis goes into some detail about measurements that he made on the relatively large changes in RI that can occur when immersion oils are use at other than room temp.

 

The bottom line is that it can be very finicky: even one deg C can make a noticeable difference. In addition,  once the oil has the WRONG RI, then the amount of spherical aberration produced will depend on the free working distance (i.e., it may look better or worse depending on the depth that you are focused into the specimen).

 

The solution as the Agard/Sedat group found years ago is to always include sub-resolution beads in your specimen and focus up-and-down on these in widefield to make sure that the point image becomes blurry in the same way, whether you focus up or down from best focus (i.e., blurry rings in both direction, not rings in one direction and blobs in the other).

 

Agard and Sedat were early users of Cargille oils mixed to the specific RI to give best results under the actual imaging conditions.

 

If trying to do the same with a water objective looking into a specimen with a higher RI than that of water, you can adjust the RI of the coupling liquid to be the same as that of the specimen with mixtures of water and glycerol, but then you will have to use the correction collar (and coverslip thickness) to compensate for the fact that you are no longer just looking into water.

 

Cheers,

 

Jim P.

 

              **********************************************
Prof. James B. Pawley,                                          Ph.  608-263-3147 
Room 223, Zoology Research Building,                                  FAX  [phone]
1117 Johnson Ave., Madison, WI, 53706                                [hidden email]
3D Microscopy of Living Cells Course, June 13-25, 2009, UBC, Vancouver Canada

Info: http://www.3dcourse.ubc.ca/                Applications still being accepted

               "If it ain't diffraction, it must be statistics." Anon.

 

Hi all,

 

Using immersion oil at 37oC.

 

Zeiss sell a range of low fluorescence immersion oils: bottles of 20, 250 and 500 ml, unfortunately all filled with the same 518F oil. According to Zeiss it's apparently 'compatible' with our Zeiss microscopes, but little extra information is offered on the Zeiss website.

 

The only info I can gleam from the web [other than the ubiquitous 518F safety data Sheet] is the following

Immersion oil "Immersol" 518 F fluorescence free, (ISO 8036-1/2, n(e)=1.518 (23°C), halogen-free)

 

So not much comment about 37oC use.  PeCon's site [who supply most of Zeiss's decent incubator stuff] have no opinion on the matter either.

 

The 518F oil safety sheet mentions "Use of the Substance / Preparation: For application information, consult the processing instructions. Processing instructions or technical

information sheet available on request.". I'll ask our knowledgeable local Zeiss Confocal rep for more details, as we are just installing a Zeiss/PeCon 37oC live cell incubator here on our Zeiss 510 MetaHead confocal.

Cargilee's site in comparison is passionate about all things immersion oil:

http://www.cargille.com/immeroil.shtml
http://www.cargille.com/immeroilselection.shtml
and their excellent, if elderly, 'primer'
http://www.cargille.com/immersionoilmicroscope.shtml

From postings to this server back in 2006, and Cargille's site not mentioning 'non-fluorescence' for type 37, it appears that Cargille 37 probably autofluoresces, so it's likely better for live cell phase-contrast [halogen lamp] transmission microscopy [but then often so are dry objectives]. I'll stick with standard 518F immersion oil at 37oC though, it's always imaged well enough on Zeiss microscopes at that temperature [and at least at 37oC it never became cloudy due to crystal precipitation]. Similarly the Cargille DF and FF seem fine to me at 37oC, and we stick ruthlessly with the same brand on each microscope. A lot of our live cell work is/was done with low power [20x Phase] air objectives though.

I do notice that immersion oil often ingresses into oil immersion objective internal optics with time, although strangely it rarely notices when imaging [like dirt on much of the internal optics, you must mostly focus through it]. One microscopy core manager even commented to me that he considers oil objectives 'consumables'. The cost of 'repair' [often the same price as a new objective] is so high we frequently have to live with it [I advise users that our say 40x objective image quality is suspect for this reason]. In truth we don't notice any drop in image quality when it must have happened, although you can see it 'smeared' on the optics when looking through the removed objective, using a magnifying glass, bright room light and a 'cleaned' objective top lens.

Old listerver posting on Cargille 37 attached below.

Regards

Keith

PS missed this link on the last post:

Dissolving crystals in Zeiss 518F oil
http://www.zeiss.com/C1256F8500454979/0/E8CD09DFA520787FC1256F86004B5FFB/$file/immersol-crystals.pdf

 

---------------------------------------------------------------------------
Dr Keith J. Morris,
Molecular Cytogenetics and Microscopy Core,
Laboratory 00/069 and 00/070,
The Wellcome Trust Centre for Human Genetics,
Roosevelt Drive,
Oxford  OX3 7BN,
United Kingdom.

Telephone:  [phone]
Email:  [hidden email]
Web-pages: http://www.well.ox.ac.uk/cytogenetics/

 

Subject:

Re: Fluorescence from Type 37 Immersion Oil

From:

Vitaly Boyko <[hidden email]>

Reply-To:

Vitaly Boyko <[hidden email]>

Date:

Wed, 16 Aug 2006 10:23:20 -0400

Content-Type:

text/plain

 

Search the CONFOCAL archive at

http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

 

PS What would be more appropriate to compare is Zeiss 518F (not 518C!) with

the Nikon type A or Cargille type DF.

 

 

 

Hi Jason,

 

I wonder why you have tested the Cargille FF imm. oil, as the refractive

index of FF oil is too far off for the standard glass (1.518) ?????!!!!!

 

Actually, I haven't noticed the "very bad" autofluorescence of the Cargille

type DF versus Zeiss 518 in the CFP, YFP or FRET channels. The difference is

less than 10%.

 

The problem with the Nikon oil is its dispenser and/or viscosity - it is

often annoying "fighting" with the air bubbles.

 

What is about the Invitrogen/Mol.Probes mounting media autofluorescence?

 

I am fed up with the irreproducibility of the background autofluorescence

using the ProlongGold mounting media!!! That is a serious problem compared

to the issue of the immersion oils.

 

Cheers,

 

Vitaly

 

NCI-Frederick

 

 

----- Original Message -----

From: "Kilgore, Jason" <[hidden email]>

To: <[hidden email]>

Sent: Tuesday, August 15, 2006 7:26 PM

Subject: Re: Fluorescence from Type 37 Immersion Oil

 

 

> Search the CONFOCAL archive at

> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

> 

> Back in 2000 I performed a comparison of four different immersion oils

> (Zeiss 518C, Nikon Type A, Cargille Type DF, and Cargille Type FF) for

> autofluorescence (using DAPI, FITC, acridine orange, and TRITC filter

> sets) and general resolution at 60x on a Nikon E400.  I didn't test

> Cargille 37.

> 

> Green autofluorescence tended to be worst for all four, but was least

> for Cargille FF, followed by Nikon Type A, then Zeiss, then (very bad)

> Cargille DF.  However, the resolution was unacceptable for Cargille FF.

> Autofluorescence in the other channels was moderate (DAPI, TRITC) to

> very low (acridine orange), with the same comparative trend applying.

> 

> Thus, ever since we have been using Nikon Type A with no complaints for

> most of our imaging.

> 

> Feel free to contact me personally and I can send you the data.

> 

> Jason

> 

> Jason A. Kilgore

> Cell Biology / Histology

> Molecular Probes/Invitrogen

> [hidden email]

> 

> 

> -----Original Message-----

> From: Confocal Microscopy List [mailto:[hidden email]] On

> Behalf Of S. Brunet

> Sent: Tuesday, August 15, 2006 11:19 AM

> To: [hidden email]

> Subject: Fluorescence from Type 37 Immersion Oil

> 

> Search the CONFOCAL archive at

> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

> 

> Hello to all,

> 

> We recently began using the Cargille 37 oil with the temperature chamber

> on the

> microscope.  I found significant fluorescence from the oil when exposed

> to the

> HBO lamp (I could not see the eYFP because of the blue background).

> 

> So I used the 20x air objective and I was able to obtain an image from

> the

> fluorescence from a drop of 37 oil placed on a coverslip as a sample

> (excitation at 488 or 458nm and pinhole at the usual confocal setting).

> 

> Is this the norm?

> 

> Regards,

> Sophie

> _______________________________________

> Sophie M.K. Brunet, Ph.D.

> Research Officer

> Optical Spectroscopist, Laser Systems and Applications

> [hidden email]

> 306-966-1719 (office)   306-966-1702 (fax)

> _______________________________________

> Saskatchewan Structural Sciences Centre

> University of Saskatchewan

> Thorvaldson Bldg.

> 110 Science Place

> Saskatoon, SK   S7N 5C9

> 

 

 

 

--