Clearing reagents with water dipping objective

[Moulding, Dale]

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Dear List,
I am looking for some advice regarding using a water dipping objective without a coverslip
and immersing directly in clearing solutions.
We have a Zeiss 20x dipper NA1.0 with correction collar (421452-9700-000).
Is this suitable to dip directly into Scalea2 and its derivatives? Is there any risk of damaging
the objective?
Cheers
Dale

[Craig Brideau]

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Most of the clearing solutions/index-matching liquids will wreck any
objective they come in contact with. The solvents used in these are pretty
nasty.
The only exceptions I can think of are the sugar-based techniques such as
SeeDB. These use fructose etc for the index-matching solution so they don't
dissolve the cement holding your lens together!

Craig

On Mon, Feb 8, 2016 at 2:36 AM, Dale Moulding <[hidden email]> wrote:

> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
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> *****
>
> Dear List,
> I am looking for some advice regarding using a water dipping objective
> without a coverslip
> and immersing directly in clearing solutions.
> We have a Zeiss 20x dipper NA1.0 with correction collar (421452-9700-000).
> Is this suitable to dip directly into Scalea2 and its derivatives? Is
> there any risk of damaging
> the objective?
> Cheers
> Dale
>

[Zdenek Svindrych-2]

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We've been told that water-based solutions are generally safe to use with
dipping lenses (I would not be afraid to use clearing solutions based on
glycerol, TDE, urea, histodenz, DMSO, fructose, focus clear, ...). Formamide
may corrode the metal parts of the lens and organic solvents (BABB, DBE,
THF, Methyl salicylate ...) may dissolve the lens cement.
Best, zdenek

--
Zdenek Svindrych, Ph.D.
W.M. Keck Center for Cellular Imaging (PLSB 003)
University of Virginia, Charlottesville, VA
http://www.kcci.virginia.edu/
tel: [phone]


---------- Původní zpráva ----------
Od: Craig Brideau <[hidden email]>
Komu: [hidden email]
Datum: 8. 2. 2016 17:29:40
Předmět: Re: Clearing reagents with water dipping objective

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Most of the clearing solutions/index-matching liquids will wreck any
objective they come in contact with. The solvents used in these are pretty
nasty.
The only exceptions I can think of are the sugar-based techniques such as
SeeDB. These use fructose etc for the index-matching solution so they don't
dissolve the cement holding your lens together!

Craig

On Mon, Feb 8, 2016 at 2:36 AM, Dale Moulding <[hidden email]> wrote:

> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> Post images on http://www.imgur.com and include the link in your posting.
> *****
>
> Dear List,
> I am looking for some advice regarding using a water dipping objective
> without a coverslip
> and immersing directly in clearing solutions.
> We have a Zeiss 20x dipper NA1.0 with correction collar (421452-9700-000).
> Is this suitable to dip directly into Scalea2 and its derivatives? Is
> there any risk of damaging
> the objective?
> Cheers
> Dale
>"

[Davide Accardi]

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Dear Dale,

I would like to make a note on the lens you are planning to use.
Th ZEISS 20x/1.0 is an optimised water dipping lens with a correction collar that allows to compensate for variation of the dipping solution refractive index in a range that spans from 1.30 to 1.36. Therefore, consider that making the experiment you briefly described you will have signifiant image aberrations due to the refractive index mismatch between the clearing solution and what the lens is optimised for.

Good luck

Davide
--
Dr. Davide Accardi
Light Microscopy Facility
Max Planck Institute of Molecular Cell Biology and Genetics
Pfotenhauerstrasse 108
01307 DRESDEN
Germany

Phone: [phone]

> On 08 Feb 2016, at 10:36, Dale Moulding <[hidden email]> wrote:
>
> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> Post images on http://www.imgur.com and include the link in your posting.
> *****
>
> Dear List,
> I am looking for some advice regarding using a water dipping objective without a coverslip
> and immersing directly in clearing solutions.
> We have a Zeiss 20x dipper NA1.0 with correction collar (421452-9700-000).
> Is this suitable to dip directly into Scalea2 and its derivatives? Is there any risk of damaging
> the objective?
> Cheers
> Dale

[Tim Feinstein]

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As I recall Olympus makes a couple of super-objectives designed for
ScaleView-A2, iDISCO etc, 10x and 25x I believe.  The other manufacturers
should have something comparable by now.  None of them will be cheap.

Best,


Tim

Timothy Feinstein, Ph.D.
Research Scientist
University of Pittsburgh Department of Developmental Biology





On 2/8/16, 4:36 AM, "Confocal Microscopy List on behalf of Dale Moulding"
<[hidden email] on behalf of [hidden email]> wrote:

>*****
>To join, leave or search the confocal microscopy listserv, go to:
>http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
>Post images on http://www.imgur.com and include the link in your posting.
>*****
>
>Dear List,
>I am looking for some advice regarding using a water dipping objective
>without a coverslip
>and immersing directly in clearing solutions.
>We have a Zeiss 20x dipper NA1.0 with correction collar
>(421452-9700-000).
>Is this suitable to dip directly into Scalea2 and its derivatives? Is
>there any risk of damaging
>the objective?
>Cheers
>Dale

[Moulding, Dale]

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Dear all,

many thanks for the helpful replies. I also heard back from my local Zeiss team, and they
say the water based reagents won't harm the objective, as long as it is cleaned with
plenty of water to prevent crystals forming after use. As Davide and Zeiss pointed out,
the RI of the objective is not ideal, but to quote from Zeiss "it's worth a try". Surely using
an objective with correction to 1.36 (for the 1.38 Scalea2) is better than a dry objective?
The correction collar has gradations past the 1.36 mark, I wonder if that gives further
correction?
There are of course specialised objectives we could use, but they are indeed very
expensive, so as a first try we will give the water dipper a go.

cheers

Dale

On Tue, 9 Feb 2016 13:40:22 +0000, Feinstein, Timothy N <[hidden email]> wrote:

>*****
>To join, leave or search the confocal microscopy listserv, go to:
>http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
>Post images on http://www.imgur.com and include the link in your posting.
>*****
>
>As I recall Olympus makes a couple of super-objectives designed for
>ScaleView-A2, iDISCO etc, 10x and 25x I believe.  The other manufacturers
>should have something comparable by now.  None of them will be cheap.
>
>Best,
>
>
>Tim
>
>Timothy Feinstein, Ph.D.
>Research Scientist
>University of Pittsburgh Department of Developmental Biology
>
>
>
>
>
>On 2/8/16, 4:36 AM, "Confocal Microscopy List on behalf of Dale Moulding"
><[hidden email] on behalf of [hidden email]>

wrote:

>
>>*****
>>To join, leave or search the confocal microscopy listserv, go to:
>>http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
>>Post images on http://www.imgur.com and include the link in your posting.
>>*****
>>
>>Dear List,
>>I am looking for some advice regarding using a water dipping objective
>>without a coverslip
>>and immersing directly in clearing solutions.
>>We have a Zeiss 20x dipper NA1.0 with correction collar
>>(421452-9700-000).
>>Is this suitable to dip directly into Scalea2 and its derivatives? Is
>>there any risk of damaging
>>the objective?
>>Cheers
>>Dale