I've 1ml anti-mouse IgE conjugated with HRP, I'll use in Elisa testing, how can I make a diluent suitable for use and what is the most perfect dilution ratio to be used?
You may use 1% BSA in PBS-T as a diluent for HRP-conjugated secondary antibody. As far as secondary antibody dilution is concerned, it would be better if you could determine the optimum dilution using a titration assay. Usually 1:2000, 1:5000 or 1:10000 dilution of secondary antibody is used for ELISA.
Hossam Ashour BSA acts as a carrier protein in the diluent. I would recommend the use IgG-free BSA which could help alleviate some of the common immunoassay problems.