Quenching glutaraldehyde autofluorescence
Nicholas King (nickk@med.su.oz.au)
Wed, 3 May 1995 17:01:07 +1000
For what it is worth, although I have never come across a satisfactory
fluorescence quencher for glutaraldehyde (and thus avoid it except when I
need to do EM), I have found that paraformaldehyde, carefully titrated
(usually between 0.5% and 4%), will often preserve antigenicity (i.e., Ab
detectability) where glutaraldehyde does not. Failing that, pure -20oC
acetone is even better at preserving Ab detectability with low background,
and it permeablises the cell into the bargain. (Naturally) there is a
penalty for low (<2%) paraformaldehyde concentration, even more so with
acetone fixation, and that is poor preservation of cellular morphology, but
it depends what you need to see - in a relatively pure population of
cardiomyocytes with high-percentage positive labelling, morphology may not
be that important. We have found both methods useful for intracellular Ab
labelling of viral antigens.
I realise this parries the question, but I hope it is helpful.
Regards,
Nicholas King
Department of Pathology
Blackburn Building, D06
University of Sydney
New South Wales 2006
Australia.
nickk@blackburn.med.su.oz.au
ph.61 2 692 4553
FAX: 61 2 692 3429
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