BABB and Methyl Salicylate

[Romin, Yevgeniy/Sloan Kettering Institute]

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Dear Listers

Our lab is working on clearing thick sections for confocal imaging and we have had great success with BABB as well as Methyl Salicylate.  We are currently looking for a way to seal a tissue submerged in these reagents underneath a coverslip to make it easier to image, since both of these reagents are very corrosive and could be harmful to the optics as well as the user.  We haven't been able to successfully seal these slides.  Has anybody here had any successful experience with sealing slides with these reagents?  It would need to be something that can resist the corrosive nature of these reagents.

Thanks to all of you in advance,

Yevgeniy
 
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[Peter Humphreys]

Not quite what you asked but.. I've used an Attofluor chamber on an invert.

Peter

Imaging Facility
WT-MRC SCI

----- Reply message -----
From: "Yevgeniy Romin" <[hidden email]>
To: <[hidden email]>
Subject: BABB and Methyl Salicylate
Date: Mon, Jan 14, 2013 8:04 pm


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Dear Listers

Our lab is working on clearing thick sections for confocal imaging and we have had great success with BABB as well as Methyl Salicylate.  We are currently looking for a way to seal a tissue submerged in these reagents underneath a coverslip to make it easier to image, since both of these reagents are very corrosive and could be harmful to the optics as well as the user.  We haven't been able to successfully seal these slides.  Has anybody here had any successful experience with sealing slides with these reagents?  It would need to be something that can resist the corrosive nature of these reagents.

Thanks to all of you in advance,

Yevgeniy
 
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[mcammer]

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Here are two ways that we have looked at large methyl salicylate whole mounts using an inverted microscope.
1.  Put in MatTek coverglass bottom dishes.  
2.  Bridge a microscope stage plate with a small hole with a large coverslip.  Put a thin line of petroleum jelly or thick silicon equivalent around hole and press coverslip down to make a seal.  Put tissue on coverslip immediately above hole.  
In both cases, a slight weight may be put on the tissue to make it flatter without squishing too much.
Here's an example from circa 2000 using a BioRad Radiance 2000:  http://www.flickr.com/photos/mcammer/3759826359/
Regards,
Michael
________________________________________________________
Michael Cammer, Assistant Research Scientist
Skirball Institute of Biomolecular Medicine
Lab: [phone]  Cell: [phone]

-----Original Message-----
From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Yevgeniy Romin
Sent: Monday, January 14, 2013 3:04 PM
To: [hidden email]
Subject: BABB and Methyl Salicylate

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Dear Listers

Our lab is working on clearing thick sections for confocal imaging and we have had great success with BABB as well as Methyl Salicylate.  We are currently looking for a way to seal a tissue submerged in these reagents underneath a coverslip to make it easier to image, since both of these reagents are very corrosive and could be harmful to the optics as well as the user.  We haven't been able to successfully seal these slides.  Has anybody here had any successful experience with sealing slides with these reagents?  It would need to be something that can resist the corrosive nature of these reagents.

Thanks to all of you in advance,

Yevgeniy
 
     =====================================================================
     
     Please note that this e-mail and any files transmitted from
     Memorial Sloan-Kettering Cancer Center may be privileged, confidential,
     and protected from disclosure under applicable law. If the reader of
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     responsible for delivering this message to the intended recipient,
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[leoncio vergara]

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Robert Zucker Cytometry Part A 69A:1143-1152 (2006). They use permount (fisher) to seal samples mounted in BABB. They use deep depression glass slides or custom made stainless steel slides to mount the tissues.

We use the stainless steel slide idea and works very well. You can easily make the chamber by drilling openings in stainless steel 1x3" plate and sealing top and bottom with coverslips and permount. Stainless steel washers may work just as well.

Leoncio Vergara

-----Original Message-----
From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Cammer, Michael
Sent: Monday, January 14, 2013 2:42 PM
To: [hidden email]
Subject: Re: BABB and Methyl Salicylate

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Here are two ways that we have looked at large methyl salicylate whole mounts using an inverted microscope.
1.  Put in MatTek coverglass bottom dishes.  
2.  Bridge a microscope stage plate with a small hole with a large coverslip.  Put a thin line of petroleum jelly or thick silicon equivalent around hole and press coverslip down to make a seal.  Put tissue on coverslip immediately above hole.  
In both cases, a slight weight may be put on the tissue to make it flatter without squishing too much.
Here's an example from circa 2000 using a BioRad Radiance 2000:  http://www.flickr.com/photos/mcammer/3759826359/
Regards,
Michael
________________________________________________________
Michael Cammer, Assistant Research Scientist Skirball Institute of Biomolecular Medicine
Lab: [phone]  Cell: [phone]

-----Original Message-----
From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Yevgeniy Romin
Sent: Monday, January 14, 2013 3:04 PM
To: [hidden email]
Subject: BABB and Methyl Salicylate

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Dear Listers

Our lab is working on clearing thick sections for confocal imaging and we have had great success with BABB as well as Methyl Salicylate.  We are currently looking for a way to seal a tissue submerged in these reagents underneath a coverslip to make it easier to image, since both of these reagents are very corrosive and could be harmful to the optics as well as the user.  We haven't been able to successfully seal these slides.  Has anybody here had any successful experience with sealing slides with these reagents?  It would need to be something that can resist the corrosive nature of these reagents.

Thanks to all of you in advance,

Yevgeniy
 
     =====================================================================
     
     Please note that this e-mail and any files transmitted from
     Memorial Sloan-Kettering Cancer Center may be privileged, confidential,
     and protected from disclosure under applicable law. If the reader of
     this message is not the intended recipient, or an employee or agent
     responsible for delivering this message to the intended recipient,
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[Glen MacDonald-2]

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Hello Yevgeniy,
Chambers for using BB+MS on inverted microscopes can be made by gluing a coverslip onto a metal support using silicone aquarium cement.  i've used  aluminum membrane slides employed with Leica laser micro dissection systems.  A thick sample can be acommodated by gluing 2 or 3 of these aluminum slides into a stack.  I request users of our LMD to toss their used membrane slides into a recycle bucket so they can be repurposed.  
Email me off line with a mailing address, I may have some extras lying around.  

Glen MacDonald
        Core for Communication Research
Virginia Merrill Bloedel Hearing Research Center
        Cellular Morphology Core
Center on Human Development and Disability
Box 357923
University of Washington
Seattle, WA 98195-7923  USA
[phone]
[hidden email]
[hidden email]





On Jan 14, 2013, at 12:04 PM, Yevgeniy Romin <[hidden email]> wrote:

> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> *****
>
> Dear Listers
>
> Our lab is working on clearing thick sections for confocal imaging and we have had great success with BABB as well as Methyl Salicylate.  We are currently looking for a way to seal a tissue submerged in these reagents underneath a coverslip to make it easier to image, since both of these reagents are very corrosive and could be harmful to the optics as well as the user.  We haven't been able to successfully seal these slides.  Has anybody here had any successful experience with sealing slides with these reagents?  It would need to be something that can resist the corrosive nature of these reagents.
>
> Thanks to all of you in advance,
>
> Yevgeniy
>
>     =====================================================================
>
>     Please note that this e-mail and any files transmitted from
>     Memorial Sloan-Kettering Cancer Center may be privileged, confidential,
>     and protected from disclosure under applicable law. If the reader of
>     this message is not the intended recipient, or an employee or agent
>     responsible for delivering this message to the intended recipient,
>     you are hereby notified that any reading, dissemination, distribution,
>     copying, or other use of this communication or any of its attachments
>     is strictly prohibited.  If you have received this communication in
>     error, please notify the sender immediately by replying to this message
>     and deleting this message, any attachments, and all copies and backups
>     from your computer.

[daj1u06]

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Not testedfor methyl salicyate but certainly for TDE and BAAB cleared samples
I use rings cut from "Stix2 double sided polyester ultra sticky clear tape"
(available from craft suppliers in UK and USA) - I have slides of cleared, whole
mount marine isopods that are over 18 months old and all are still good - the
tape is about 125um thick and lives up to its name, it is incredibly sticky -  
grips stronger than a politician holding onto power :-).

I cut a length off, relocate on roll, cut off the double thicknes strip (backing,
tape, backing, tape) , throw away the exposed tape and then use a wad
punch set to cut the holes out to whatever size I want.

If we want to image both sides or are not sure of the orientation of samples,
we often mount between 2 large coverslips using a ring of this stuff.