Flash Frysning och Cryosectioning E12.5 Mouse Brain (Translated to Swedish)

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Cite this Article: Flash Frysning och Cryosectioning E12.5 Mouse Brain

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 Frysning och Cryosectioning E12.5 Mouse Brain

Fix vävnad i 4% paraformaldehyd i PBS för önskad tid.
Sackaros ingjuta vävnad (cryoprotection)
1. Gör 30% sackaroslösning i PBS w / v under 2059 rör.
2. Skölj vävnad 3x i PBS (~ 5 min med gungande).
3. Placera vävnad i 30% sackaroslösning. Tissue kommer inte att sjunka.
4. Placera vävnad i 4 ° C över natten, eller tills den har sjunkit.
Etikett lämplig storlek cryomold med information och orientering.
Fyll cryomold med oktober (undvik bubblor).
Överföring vävnad till oktober bad och päls den med oktober
Överföring vävnad till oktober i cryomold.
Orient vävnaden i mikroskop.
Häll flytande kväve i plast petriskål.
Snabbt och försiktigt sänka vävnad i cryomold till kväve. (Inte doppa toppen av cryomold.)
När oktober är solid vit, placera fryst vävnad till -80 ° C frys för förvaring.
Jämvikta vävnad till ~ 20 ° C under minst 30 min. innan snittning.

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Disclosures: Flash Frysning och Cryosectioning E12.5 Mouse Brain

Materials: Flash Frysning och Cryosectioning E12.5 Mouse Brain

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 Frysning och Cryosectioning E12.5 Mouse Brain

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

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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 Frysning och Cryosectioning E12.5 Mouse Brain