Best dipping lenses?

[Craig Brideau]

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

We are looking for a reliable dipping lens for confocal and two photon
microscopy work.  We have used a 60x 1NA Nikon in the past but we have
had problems with moisture.  I would like to know what sorts lenses
people are using (Nikon/Zeiss/Olympus/...) and how reliable and
effective they are.  What are you using, does it give good results,
and how reliable is it?

[Moninger, Thomas]

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

I have had good Confocal and MP results with Nikon's Fluor 20x/0.50W and
40x/0.8W dipping lenses. They have been used for both cell culture and
in-vivo imaging. Tom

-----Original Message-----
From: Confocal Microscopy List [mailto:[hidden email]] On
Behalf Of Craig Brideau
Sent: Wednesday, September 05, 2007 1:52 PM
To: [hidden email]
Subject: (SCL: 6) Best dipping lenses?


Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

We are looking for a reliable dipping lens for confocal and two photon
microscopy work.  We have used a 60x 1NA Nikon in the past but we have
had problems with moisture.  I would like to know what sorts lenses
people are using (Nikon/Zeiss/Olympus/...) and how reliable and
effective they are.  What are you using, does it give good results,
and how reliable is it?

[Craig Brideau]

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

Thanks for the reply!  Have you ever had any problems with the Nikon
objectives?  How did you find Nikon's customer service/support
overall?

Craig Brideau


On 9/5/07, Moninger, Thomas <[hidden email]> wrote:

> Search the CONFOCAL archive at
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
> I have had good Confocal and MP results with Nikon's Fluor 20x/0.50W and
> 40x/0.8W dipping lenses. They have been used for both cell culture and
> in-vivo imaging. Tom
>
> -----Original Message-----
> From: Confocal Microscopy List [mailto:[hidden email]] On
> Behalf Of Craig Brideau
> Sent: Wednesday, September 05, 2007 1:52 PM
> To: [hidden email]
> Subject: (SCL: 6) Best dipping lenses?
>
>
> Search the CONFOCAL archive at
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
> We are looking for a reliable dipping lens for confocal and two photon
> microscopy work.  We have used a 60x 1NA Nikon in the past but we have
> had problems with moisture.  I would like to know what sorts lenses
> people are using (Nikon/Zeiss/Olympus/...) and how reliable and
> effective they are.  What are you using, does it give good results,
> and how reliable is it?
>

[Julio Vazquez]

Search the CONFOCAL archive at http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

-

Craig, 

If Zeiss is an option, we have the Zeiss C-Achroplan NIR 40/0.8 W. We actually use it as a long-working distance immersion lens on an inverted scope... not super-convenient because of the large volume of water required, but it gives much better images than a conventional long-working distance lens. Works quite well  for two-photon, as it is a relatively new lens designed for high transmission in the NIR. I would assume it would work very well in its intended normal use as a dipping lens. We had for a while a Zeiss Achroplan 60x/0.9 (If I remember correctly), but we returned it since it was redundant with the 40/0.8, and although a good lens, we thought the C-Achro 40/0.8 was a bit better in terms of image quality.

--

Julio Vazquez, 

Fred Hutchinson Cancer Research Center

Seattle, WA 98109-1024

[hidden email]

http://www.fhcrc.org/science/shared_resources/imaging/

On Sep 5, 2007, at 11:51 AM, Craig Brideau wrote:

Search the CONFOCAL archive at

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

We are looking for a reliable dipping lens for confocal and two photon

microscopy work.  We have used a 60x 1NA Nikon in the past but we have

had problems with moisture.  I would like to know what sorts lenses

people are using (Nikon/Zeiss/Olympus/...) and how reliable and

effective they are.  What are you using, does it give good results,

and how reliable is it?

[Craig Brideau]

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

Thanks for the tip!  This sounds like an interesting lens.  Have you
used Zeiss objectives for a while?  How do you find their reliability,
and have you ever had to send one for repair?

Craig


On 9/5/07, Julio Vazquez <[hidden email]> wrote:

> Search the CONFOCAL archive at
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
> -
> Craig,
>
> If Zeiss is an option, we have the Zeiss C-Achroplan NIR 40/0.8 W. We
> actually use it as a long-working distance immersion lens on an inverted
> scope... not super-convenient because of the large volume of water required,
> but it gives much better images than a conventional long-working distance
> lens. Works quite well  for two-photon, as it is a relatively new lens
> designed for high transmission in the NIR. I would assume it would work very
> well in its intended normal use as a dipping lens. We had for a while a
> Zeiss Achroplan 60x/0.9 (If I remember correctly), but we returned it since
> it was redundant with the 40/0.8, and although a good lens, we thought the
> C-Achro 40/0.8 was a bit better in terms of image quality.
>
> --
> Julio Vazquez,
> Fred Hutchinson Cancer Research Center
> Seattle, WA 98109-1024
>
> [hidden email]
> http://www.fhcrc.org/science/shared_resources/imaging/
>
>
>
>
>
> On Sep 5, 2007, at 11:51 AM, Craig Brideau wrote:
>
> Search the CONFOCAL archive at
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
> We are looking for a reliable dipping lens for confocal and two photon
> microscopy work.  We have used a 60x 1NA Nikon in the past but we have
> had problems with moisture.  I would like to know what sorts lenses
> people are using (Nikon/Zeiss/Olympus/...) and how reliable and
> effective they are.  What are you using, does it give good results,
> and how reliable is it?
>

[Stephen Cody]

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

G'day Graig,

If I may answer a question you haven't asked!....

Before making a decision on a dipping lens (presumably you are going to
purchase a whole new confocal / multiphoton to go with that lens), you
should ask all the manufactures to report what percentage of the back
focal plane of the dipping lens is filled. Ask for a report from the
factory (don't rely on a response of the sales team on this one, they
are not likely to be aware). Also let them know that it is easy enough
for you to test, that way you will receive a frank response.

Many of these High NA, Low magnification dipping lenses are drastically
under filled. Hence they are only operating at a fraction of the
nominated NA unless installed on the physiology microscopes for which
they were developed. Unless we ask the manufacturers to do something
about that nothing will change.

If a 20x dipping lens and a 60x dipping lens are marked with the same
NA, then the 20X lens used with an optical zoom of 3 should perform
similarly to the 60x lens at a zoom of 1. This would be of benefit for
live cell experiments as you could use low power to find the cells of
interest and then simply zoom in for high resolution images.

However, I believe in most installations the high NA 20x dipping lenses
are under filled and so this strategy does not usually work.

Cheers

Stephen H. Cody
Microscopy Manager
Central Resource for Advanced Microscopy
Ludwig Institute for Cancer Research
PO Box 2008 Royal Melbourne Hospital
Parkville, Victoria,      3050
Australia
Tel: [phone]    Fax: [phone]
email: [hidden email]
www.ludwig.edu.au/labs/confocal.html
www.ludwig.edu.au/confocal



This communication is intended only for the named recipient and may contain information that is confidential, legally privileged or subject to copyright; the Ludwig Institute for Cancer Research does not waiver any rights if you have received this communication in error.
The views expressed in this communication are those of the sender and do not necessarily reflect the views of the Ludwig Institute for Cancer Research.

[Jacqueline Ross]

Search the CONFOCAL archive at http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

Hi Julio,

 

Just wondering what your method of maintaining the water column is for the long working distance water immersion objective on an inverted microscope? I’ve heard various suggestions before such as glove fingers, etc. Do you do something similar?

 

Kind regards,

 

Jacqui

 

Jacqueline Ross
Biomedical Imaging Research Unit 
School of Medical Sciences 
Faculty of Medical & Health Sciences
The University of Auckland
Private Bag 92019
Auckland, NEW ZEALAND

Tel: [phone] Ext 87438
Fax: [phone]

http://www.health.auckland.ac.nz/biru/

---

From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Julio Vazquez
Sent: 06 September 2007 08:46
To: [hidden email]
Subject: Re: Best dipping lenses?

 

-

Craig, 

 

If Zeiss is an option, we have the Zeiss C-Achroplan NIR 40/0.8 W. We actually use it as a long-working distance immersion lens on an inverted scope... not super-convenient because of the large volume of water required, but it gives much better images than a conventional long-working distance lens. Works quite well  for two-photon, as it is a relatively new lens designed for high transmission in the NIR. I would assume it would work very well in its intended normal use as a dipping lens. We had for a while a Zeiss Achroplan 60x/0.9 (If I remember correctly), but we returned it since it was redundant with the 40/0.8, and although a good lens, we thought the C-Achro 40/0.8 was a bit better in terms of image quality.

 

--

Julio Vazquez, 

Fred Hutchinson Cancer Research Center

Seattle, WA 98109-1024

 

[hidden email]

http://www.fhcrc.org/science/shared_resources/imaging/

 

 

 

On Sep 5, 2007, at 11:51 AM, Craig Brideau wrote:

Search the CONFOCAL archive at

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

 

We are looking for a reliable dipping lens for confocal and two photon

microscopy work.  We have used a 60x 1NA Nikon in the past but we have

had problems with moisture.  I would like to know what sorts lenses

people are using (Nikon/Zeiss/Olympus/...) and how reliable and

effective they are.  What are you using, does it give good results,

and how reliable is it?

 

[Craig Brideau]

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

That's a great tip!  Thanks!  We actually have one completly homebrew
rig, and two heavily modifed Nikons for multiphoton.  We use our own
telescopes and scan lenses at the input port to size our incident beam
such that the objective apertures are filled.  I had no idea that it
was such a problem in off-the-shelf units though!

Craig


On 9/5/07, Stephen Cody <[hidden email]> wrote:

> Search the CONFOCAL archive at
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
> G'day Graig,
>
> If I may answer a question you haven't asked!....
>
> Before making a decision on a dipping lens (presumably you are going to
> purchase a whole new confocal / multiphoton to go with that lens), you
> should ask all the manufactures to report what percentage of the back
> focal plane of the dipping lens is filled. Ask for a report from the
> factory (don't rely on a response of the sales team on this one, they
> are not likely to be aware). Also let them know that it is easy enough
> for you to test, that way you will receive a frank response.
>
> Many of these High NA, Low magnification dipping lenses are drastically
> under filled. Hence they are only operating at a fraction of the
> nominated NA unless installed on the physiology microscopes for which
> they were developed. Unless we ask the manufacturers to do something
> about that nothing will change.
>
> If a 20x dipping lens and a 60x dipping lens are marked with the same
> NA, then the 20X lens used with an optical zoom of 3 should perform
> similarly to the 60x lens at a zoom of 1. This would be of benefit for
> live cell experiments as you could use low power to find the cells of
> interest and then simply zoom in for high resolution images.
>
> However, I believe in most installations the high NA 20x dipping lenses
> are under filled and so this strategy does not usually work.
>
> Cheers
>
> Stephen H. Cody
> Microscopy Manager
> Central Resource for Advanced Microscopy
> Ludwig Institute for Cancer Research
> PO Box 2008 Royal Melbourne Hospital
> Parkville, Victoria,      3050
> Australia
> Tel: [phone]    Fax: [phone]
> email: [hidden email]
> www.ludwig.edu.au/labs/confocal.html
> www.ludwig.edu.au/confocal
>
>
>
> This communication is intended only for the named recipient and may contain information that is confidential, legally privileged or subject to copyright; the Ludwig Institute for Cancer Research does not waiver any rights if you have received this communication in error.
> The views expressed in this communication are those of the sender and do not necessarily reflect the views of the Ludwig Institute for Cancer Research.
>
>

[Julio Vazquez]

Search the CONFOCAL archive at http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

-

Hi Jacqui, 

Our 40x/0.8 Water lens has a working distance of 1.75 mm. This means that if we focus on a sample that is mounted right near the coverslip, we would have a 1.75 mm bubble/column of water to maintain between lens and coverslip. Keeping that rather large bubble of water from running down can be tricky, but generally that is not how we use the lens. We use the lens either for samples mounted close to the coverslip, but for which we need more than the approximately 100-150 microns of working distance of high NA oil/water immersion lenses (but typically, not more than 400 microns), or when we try to image "weird" samples where the desired focal plane is well past the coverslip (or deep inside a well, etc...)

 When we purchased our particular lens, it came with a rubber sleeve that wraps around the collar of the lens (the objective tip itself is rather narrow and made of a teflon-like material), and extends slightly past the tip of the lens, a bit like a lens cap with a hole. This sleeve can be moved within a certain range. This effectively reduces the working distance, but it is generally not a problem since for most samples we don't usually need more than 0.3-0.5 mm of working distance. Therefore, we adjust the rubber sleeve to allow for about 0.5-1.0 mm working distance, and it is generally no problem to maintain a water bubble of this size. In the second situation, the distance between lens and coverslip will also be substantially less than 1.75 mm, since we are using the WD to reach deep inside the sample. If you need to image a sample right from the coverslip to over 1 mm deep, then you need a large column of water and some means to keep it from spilling, while not interfering with the focusing. In that situation, you may need one of those systems with a flexible latex sleeve that can hold more water while adapting to the focusing distance, but we never had a need for that. I believe there was a thread on this on the listserver some time ago, so you may be able to find some extra tips there. 

If you have a Zeiss lens, you should be able to get one of those rubber caps from your rep; otherwise, they should be easy to manufacture from standard rubber/plastic tubing. Something that may work too is to use some of the teflon tape normally used to seal joints... very thin and flexible: just wrap some at the tip of the lens, so that it extends a little past the front...

--

Julio Vazquez

Fred Hutchinson Cancer Research Center

Seattle, WA 98109-1024

[hidden email]

http://www.fhcrc.org/science/shared_resources/imaging/

On Sep 6, 2007, at 5:52 PM, Jacqui Ross wrote:

Search the CONFOCAL archive at http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

Hi Julio,

 

Just wondering what your method of maintaining the water column is for the long working distance water immersion objective on an inverted microscope? I’ve heard various suggestions before such as glove fingers, etc. Do you do something similar?

 

Kind regards,

 

Jacqui

 

Jacqueline Ross
Biomedical Imaging Research Unit 
School of Medical Sciences 
Faculty of Medical & Health Sciences
The University of Auckland
Private Bag 92019
Auckland, NEW ZEALAND

Tel: [phone] Ext 87438
Fax: [phone]

http://www.health.auckland.ac.nz/biru/

---

From: Confocal Microscopy List [[hidden email]] On Behalf Of Julio Vazquez
Sent: 06 September 2007 08:46
To: [hidden email]
Subject: Re: Best dipping lenses?

 

-

Craig, 

 

If Zeiss is an option, we have the Zeiss C-Achroplan NIR 40/0.8 W. We actually use it as a long-working distance immersion lens on an inverted scope... not super-convenient because of the large volume of water required, but it gives much better images than a conventional long-working distance lens. Works quite well  for two-photon, as it is a relatively new lens designed for high transmission in the NIR. I would assume it would work very well in its intended normal use as a dipping lens. We had for a while a Zeiss Achroplan 60x/0.9 (If I remember correctly), but we returned it since it was redundant with the 40/0.8, and although a good lens, we thought the C-Achro 40/0.8 was a bit better in terms of image quality.

 

--

Julio Vazquez, 

Fred Hutchinson Cancer Research Center

Seattle, WA 98109-1024

 

[hidden email]

http://www.fhcrc.org/science/shared_resources/imaging/

 

 

 

On Sep 5, 2007, at 11:51 AM, Craig Brideau wrote:

Search the CONFOCAL archive at

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

 

We are looking for a reliable dipping lens for confocal and two photon

microscopy work.  We have used a 60x 1NA Nikon in the past but we have

had problems with moisture.  I would like to know what sorts lenses

people are using (Nikon/Zeiss/Olympus/...) and how reliable and

effective they are.  What are you using, does it give good results,

and how reliable is it?

 

[Michael Weber-4]

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

Hi Julio,

why are you not using water-immersion oil like Zeiss "Immersol W"?

cheers,
Michael


Julio Vazquez wrote:
> Our 40x/0.8 Water lens has a working distance of 1.75 mm. This means
> that if we focus on a sample that is mounted right near the coverslip,
> we would have a 1.75 mm bubble/column of water to maintain between lens
> and coverslip. Keeping that rather large bubble of water from running
> down can be tricky, but generally that is not how we use the lens.

[Julio Vazquez]

Search the CONFOCAL archive at http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal -

Hi Michael, 

we don't always use "immersol W" for a couple of not very profound reasons: 

1. water is free, while "immersol W" has to be purchased. 

2. Water seems to have higher surface tension, so for keeping a large bubble of water between objective and coverslip, water works better... immersol tends to run down more easily

3. immersol, on the other hand, seems to be more resistant to evaporation, so for high NA water objectives that require very little immersion fluid, such as the 40/1.2 W, immersol works fine, and is better  for long term experiments

so basically, if we can use water, we use water... for long-term experiments, we use immersol...

--

Julio Vazquez

Fred Hutchinson Cancer Research Center

Seattle, WA 98109-1024

http://www.fhcrc.org/

 

==

On Sep 10, 2007, at 5:13 AM, Michael Weber wrote:

Hi Julio,

why are you not using water-immersion oil like Zeiss "Immersol W"?

cheers,

Michael