sandwich ELISA

Martin Offterdinger a8803349 at unet.univie.ac.at
Wed Jan 22 05:54:35 EST 1997


Hi
I need help setting up a sandwich ELISA for a transmembrane receptor,
that shows low expression. 
I tried to precipitate the receptor from a cell lysate using a plate
bound monoclonal AB and quantitate with a polyclonal AB followed by
staining with anti rabbit HRP. At the moment I do not get a specific
signal.
My questions are :

1 Will it be helpful to use the polyclonal antibody as the
primary(coat it to the plate) ?

2 What detergent should be used and in which concentration to
solubilise the cells??-At the moment I use 0.5% digitonin in isotonic
buffer. 
Do you think that different detergents would work better???
Thank you very much 
martin





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