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HELP - (Oct/12/2006 )

Somebody how the double-capture ELISA works?


do you mean double ELISA (or sandwich ELSA)?

you coat first a capture antibody (often monoclonal)
then you block non specific binding sites with non fat dry milk, or BSA...
then you add your standard (recombinant protein) and your sample
then you add the detection antibody (often biotinylated antibody, polyclonal)
after the bioitnylated antibody, you add streptavidin-HRP, or streptavidin AP
then you add substrate, let incubate until the color is OK and you read.

Between each step (but not after adding substrate) you wash three times with wash buffer (often PBS plus tween-20)