Immunoprecipitation with protein A agarose Use lysates prepared using freon extraction add 0.5-1 µL of antibody - incubate with end-over-end rotation for 1-4 hours at 4°C. add 40µL protein-A-agarose, incubate with end-over-end rotation for 1-2 hours at 4°C. recover beads by centrifugation, at 4°C, 2 minutes at 2000 rpm. Remove lysate by aspiration --it is ok to leave some liquid above the pellet. resuspend in your original lysis buffer (1.5ml) and wash for 5 min. at 4°C (with end-over-end rotation) recover beads by centrifugation, at 4°C, 2 min. at 2000 rpm resuspend in high salt wash buffer (buffer 2) and wash for 5 min. at 4°C recover beads by centrifugation, at 4°C, 2 minutes at 2000 rpm
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