immunofluorescence: direct vs. indirect

P.C. peter.cherepanov at uz.kuleuven.ac.be
Tue Jun 23 07:15:06 EST 1998


What would be the best choice for doing immunofluorescence:
to obtain labeled monoclonal antibodies or to use 
labeled secondary antibody ?

I heard that using secondary antibodies increases sensitivity - 
is that true ? 

(As I understand the primary antibody is usually an IgG, and 
the bulk of the secondary is IgGs too, which are monomeric. 
Therefore, I would expect only a drop in sensitivity and 
specificity. Using biotinilated antibodies may have an advantage, 
since avidine is tetrameric and can bring more FITC residues to 
the spot (?))

I need not only sensitive, but also very specific detection. 
(surface staining of monocytes, I plan to use CD14 antibody).

Would like to hear any hints.

Peter



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