Immunofluorescence-sensitivity

Guy Oshiro Guy.Oshiro at UCHSC.edu
Fri Jul 25 20:33:24 EST 1997


In article <5r5jq5$9on at tali.UCHSC.edu>
Guy.Oshiro at UCHSC.edu (Guy Oshiro) writes:

> I think that you can expect about a 4-fold increase in sensitivity by
> the above approach.  Another approach that you should look into is the
> use of a fluorescent substrate for an enzyme linked reaction.  Amersham
> sells an ECF kit (enhanced Chemi Fluorescence) that basically couples
> Alk Phos to the secondary and the AP acts on a fluorescence substrate
> (called AttoPhos).  I am planning to test that particular system out in
> our lab this week - so I don't have any real data to share with you.

Oops, looks like that I goofed in this post.  I was thinking about
looking at the protein by westerns by using a fluorescent system.  But
I think that the poster was referring to IM microscopy.  So forget the
part about the AttoPhos substrate - but the 4 fold increase still
holds.  Another thing to consider is to make a tertiary antibody tree 
-- 1' against protein of interest, 2' against 1' antibody conjugated to
a flurophore, 3' against 2' also conjugated to the same fluorophore.

Guy Oshiro
EMAIL: Guy.Oshiro at UCHSC.edu

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