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fixation of tissues using 4% paraformaldehyde

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kachang
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Topic Started by kachang
on 5/23/2011 2:21 AM   
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I am fixing rat ovary using 4% paraformaldehye for IHC.tissues are not intact but nick and crack appears around the tissues  on sectioning.what exact steps and timings should i follow.pls help


Last edited May 29, 2012, 4:07 AM by pundir
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pundir
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Posted By pundir
on 5/23/2011 18:45 PM   
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 Hi,
the fixation time depends on thickness of your tissue, for rat ovary overnight would be enough also check the pH of PFA , clarify how you are processing your tissue after fixation means are you going for cryosectioning or paraffin sectioning if cryo then are you going for cryoprotection in 30 % sucrose befoere sectioning.............elaborate your querry for further help

impossible! let's do it................



kachang
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Posted By kachang
on 5/27/2011 2:01 AM   
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thank u pundir. the ovary size is about 3cm x 3cm, PFA of pH7.4, and tissue processed in paraffin.tissue fixed for 72 hours as tissue doesnt get fixed in 24 or 48 hrs.fixed tissue are processed as follows: PBS 1x wash - 3x -30 min each dehydration in alcohol series of 30%,50% -30 min each, 70% -overnight, 90%,100% -30 min each, 100%:xylene-20min xylene I & II -20 min each Xylene : wax - 45 min wax I & II - 45 min each wax III - 1 hr embedding



drgaju.15
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Posted By drgaju.15
on 2/28/2012 15:07 PM   
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hai
can any body tell me 4% paraformaldehyde and 10% formalin is same or different

thanks



Matizales
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Posted By Matizales
on 5/29/2012 0:30 AM   
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Hello,

I hope you solve your doubts.
Anyway, 4% formalin and 10% formalin is the same, as it comes from 35-37'5% and you could consider it as 4% (37%) or 10% if you add 9 times solvent.
Paraformaldehyde is the polymer and you can consider it in the same way. You will use PFA when the fixation concentration is critical.

Good luck,

Pedro

   



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