Re: [Histonet] Formalin Fixation times for IHC samples
Dear Anne:
Standard answer, it depends on the antigen in question. Some survive
fixation for 24 hours or more, some are rendered unreactive in 4 hours.
Formalin works slowly so longer fixation usually reduces
immunoreactivity. Also, the method of detection is important. In my
hands the Vector ABC Elite kit gives a much stronger signal than the
standard kit and often gives good results when the standard kit shows
nothing.
Geoff
Anne C Lewin wrote:
> Could I get a general concensus from Histoland about the amount of
> time needed to fix a sample for IHC without over-fixing? My
> training/journal searches have led me to believe that the longer a
> sample is in formalin, the stronger the protiens are cross-linked, and
> the more difficult it is to break those bonds with antigen retreival
> and get your antibody to the target. I generally tell the scientists
> in my department to fix overnight (most samples vary from about the
> size of a pea to a lima bean), and then to transfer to 70% Ethanol for
> me to process for paraffin. The total time in fixative tends to be
> around 24 hours. Morphology has always been great, and my IHC's work
> well. This question is mainly for my information, since I have used
> the same tissue prep protocol for a few years now, I want to make sure
> I am keeping up with current opinions.
> Thanks!
> -Anne
>
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Geoff McAuliffe, Ph.D.
Neuroscience and Cell Biology
Robert Wood Johnson Medical School
675 Hoes Lane, Piscataway, NJ 08854
voice: (732)-235-4583; fax: -4029
mcauliff@umdnj.edu
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