dilution+of+antibody+when+used+with+super+g+blocking+buffer

question for Grace Biolabs

Hi Grace Biolabs representative, I am curious about what I should dilute my antibody in when I am using it after the Super G Blocking Buffer has been used. I have nitrocellulose slides spotted with tumor cDNA expression library cell lysate which will be blocked with Super G. I will then probe this slide with antibody. The typical incubation buffer that we use in our lab consists of 5 mL 30% BSA, 25 uL of Tween 20, 5 mL 10X PBS, and 40 mL H20. However, I wonder if it would be best for me to avoid incubating the antibody with BSA since the blocking buffer had no BSA in it and Super G Blocking Buffer was used. In this situation, perhaps I should just dilute the antibody in PBS for best results? Or maybe PBS with some Tween 20? Thanks for any feedback you have to offer!

Grace Biolabs reply

Hi Kurt, I recently was forwarded your question regarding incubation buffer for primary antibody following Super G blocking of nitrocellulose. Normally, here for in house assays I have used PBS with 0.1% Tween20. Using BSA should not be a problem either though, and for longer incubations, may help to prevent protein aggregation. I would test both to see what performs best for your application but I do not foresee any issues with either. Please let me know if you have any further questions and I would be happy to answer them. Best Regards, Michael