In addition, can we use the 1.0mg/ml antibody for HRP-antibody conjugation? According to peroxidase labeling kit we should use 0.3ml @ 4.0mg/ml. However, this is too high of a concentration and it is difficult to manage.
i think you have to follow ELISA Guidebook chapter 4 (titration of reagents) which discuss in details the chessboard or checkerboard titration of reagents used for ELISAs and how to deal with poor results and that all are attached with this replay i hope you gonna find that helpful
Coating concentatrion: 1 ... 10 µg/mL
Antigen concentration (detectable): depends on the affinity of the primary antibody. Normally between 1 pg/mL ... 10 ng/mL.
Conjuate concentration: 0.1 ... 1 µg/mL (dilute your conjugate 1 : 1000)
us PBS 0.3 mol/L NaCl. The increased NaCl will avoid unspecific reactions.
i think you have to follow ELISA Guidebook chapter 4 (titration of reagents) which discuss in details the chessboard or checkerboard titration of reagents used for ELISAs and how to deal with poor results and that all are attached with this replay i hope you gonna find that helpful
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