Flash Invriezen en Cryosectioning E12.5 hersenen van muizen (Translated to Dutch)

You must be subscribed to JoVE to access this content.

This article is a part of JoVE General. If you think this article would be useful for your research, please recommend JoVE to your institution's librarian.

Current Access Through Your IP Address

You do not have access to any JoVE content through your current IP address.

IP: 50.17.109.248, User IP: 50.17.109.248, User IP Hex: 840003064

Current Access Through Your Registered Email Address

You aren't signed into JoVE. If your institution subscribes to JoVE, please sign in or create an account with your institutional email address to access this content.

Cite this Article: Flash Invriezen en Cryosectioning E12.5 hersenen van muizen

Currle, D. S., Monuki, E. S. Flash Freezing and Cryosectioning E12.5 Mouse Brain. J. Vis. Exp. (4), e198, doi:10.3791/198 (2007).

Protocol: Flash Invriezen en Cryosectioning E12.5 hersenen van muizen

Fix weefsel in 4% paraformaldehyde in PBS voor de gewenste tijd.
Sucrose trekken weefsel (cryoprotection)
1. Maak 30% sucrose-oplossing in PBS w / v in 2059 buis.
2. Spoel weefsel 3x in PBS (~ 5 min met schommelen).
3. Plaats weefsel in 30% sucrose oplossing. Weefsel zal niet zinken.
4. Plaats het weefsel in 4 ° C 's nachts, of totdat het is gezonken.
Label juiste grootte cryomold met informatie en oriëntatie.
Vul cryomold met oktober (vermijd luchtbellen).
Overdracht weefsel oktober bad en coaten met oktober
Overdracht weefsel LGO cryomold.
Orient het weefsel onder de microscoop.
Giet vloeibaar stikstof in plastic petrischaal.
Snel en zorgvuldig lager het weefsel in cryomold in de stikstof. (Niet onderdompelen de top van de cryomold.)
Wanneer de LGO solide is wit, plaats de bevroren weefsel in -80 ° C vriezer voor de opslag.
Evenwicht weefsel tot ~ 20 ° C gedurende ten minste 30 minuten. voorafgaand aan snijden.

Subscription Required. Please recommend JoVE to your librarian.

Disclosures: Flash Invriezen en Cryosectioning E12.5 hersenen van muizen

Materials: Flash Invriezen en Cryosectioning E12.5 hersenen van muizen

Name	Company	Catalog Number	Comments
Tissue-Tek Cryomold	Ted Pella, Inc.	27181
O.C.T.	Ted Pella, Inc.	27050
Sucrose solution			30% sucrose solution in PBS w/v
paraformaldehyde			4% paraformaldehyde in PBS

Ask the Author: Flash Invriezen en Cryosectioning E12.5 hersenen van muizen

11 Comments

This is one application almost all labs use either for some immuno or insitu experiments and seeing the procedure to this level of detail, I think will enable people to achieve better results with less trial and error.

Posted by: AnonymousJuly 6, 2007, 9:41 PM

awesome

Posted by: AnonymousJuly 18, 2007, 10:42 PM

great! thanks a lot.

Posted by: AnonymousSeptember 25, 2007, 2:01 PM

The presentation was wonderful and I learned more from this video than what my conservative collegues in the lab explained me.

Thanks a lot

Posted by: #3May 29, 2008, 4:58 PM

I'm having some issues washing off the OCT with BPS from the slides. - I did some retrograde staining of peripheral nerves and I cross sectioned the spinal cord of mice-
the problem is that as I add PBS drop by drop, or as I place the slide into plate with PBS liquid layer my samples keep falling off the slide..
Any ideas? because I'm running out of them
e-mail me please: jccs_85@hotmail.com

Posted by: CarlosJuly 2, 2009, 9:35 AM

This is a great video demonstrating how to cryosection brain that will certainly help me with my future studies. I was wondering, is it always necessary and/or desirable to fix the brain with paraformaldahyde prior to cryoprotecting with sucrose? I seem to recall hearing for some applications, such as IHC, paraformaldahyde fixation may disrupt the antibody/antigen interaction for certain antibodies used. Would someone be willing to comment on this?

Posted by: Shelly July 8, 2009, 8:18 PM

send me an e-mail at spencer.currle@stjude.com

6.1

Posted by: AnonymousJuly 9, 2009, 10:32 AM

Posted by: Shelly July 9, 2009, 4:42 PM

I have the same question as Shelly. Some applications require fixing tissue after cryosectioning. Do you have a protocol for this?

Posted by: SueSeptember 20, 2010, 3:03 PM

Hello, my name is Xiang Weng, I am a student in Long Island University, department of Biomedical Science, I want to learn cryosectioning, thank you.

Posted by: Xiang W.October 5, 2012, 11:24 AM

Hello, my name is Xiang Weng, I am a student in Long Island University, department of Biomedical Science, I want to learn cryosectioning, thank you.

Posted by: Xiang W.October 5, 2012, 11:35 AM

I have looked all over for the answer but I cannot find it: How many seconds (or how long in general) does it take to flash freeze a serum lab sample using ethanol and dried ice? Thanks!

Posted by: Michele K.April 3, 2013, 8:51 PM

Post a Question / Comment / Request

You must be signed in to post a comment. Please sign in or create an account.

Date Published	5/28/2007
JoVE Section	General
JoVE Issue	4
Comments	11 Comments
View Count	Loading... (Details…)
DOI	doi:10.3791/198

Automatic Translation

Flash Invriezen en Cryosectioning E12.5 hersenen van muizen