Re: [Histonet] RE: Antibody staining on non-pfa fixed tissue
I have had good results with mouse and rat brain fixed in 70% Alcohol.
National Institutes of Health
National Institute of Dental and Craniofacial Research
Pain and Neurosensory Mechanism Branch
On 7/31/06 3:56 AM, "C.M. van der Loos" wrote:
> Dear Greg,
> As you already concluded the epitope of interest is PFA-sensitive. How
> about using a non-crosslinking fixative here? For example methacarn
> (methanol:chloroform:acetic aced = 6:3:1) does not contain
> formaldehydes. Fix your sample for 48 hours, dehydrate via alcohol
> series and embed in paraffin. Because methacarn is non-crosslinking
> fixative most antigens doesn't need antigen retrieval.
> Just a suggestion...
> Chris van der Loos, PhD
> Dept. of Pathology
> Academic Medical Center M2-230
> Meibergdreef 9
> NL-1105 AZ Amsterdam
> The Netherlands
> Date: Fri, 28 Jul 2006 18:46:26 +0200
> From: "Gregory A. O'Sullivan"
> Subject: [Histonet] Antibody staining on non-pfa fixed tissue
> To: "HistoNet"
> Hello HistoNetters,
> I realise that this may be a question that has arisen on previous
> occasions, however, I have been unable
> to find an exact answer from a search of previous email correspondence
> on HistoNet.
> I would like to detect and examine the distribution of my protein of
> interest in mouse brain sections
> (with particular emphasis on the hippocampus) and compare it to other
> known markers.
> The problem is that the antibody I am using is extremely sensitive to
> pfa fixation but the marker antigens
> are best detected on fixed tissue. I have tried to overcome this
> problem by fixing cryostat sections
> briefly with pfa before using different antigen retrieval protocols
> (either microwave or heated bath incubatio! n
> in C
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