Do you need to dialyze cytoplasmic lysate to remove free biotin when purifying in vivo biotin labelled proteins from insect cells?

01 January 2013 1 3K Report

I am looking at biotinylating a protein in vivo in insect cells where the insect cell media will be supplemented with 4 uM biotin. The protein accumulates in the cytoplasm. The question is: has anyone purified a biotin tagged protein from insect cell lysates using Pierce Monomeric Avidin Ultralink resin (or similar resin)? And did you have to dialyze the cytoplasmic lysate to get rid of the free biotin? I wash the insect cells in PBS to get rid of media associated biotin before lysing them and would prefer not to dialyze the cytoplasmic lysate.

Nell Matthews

We purify biotin labeled proteins several ways: dialysis against avidin-containing buffer, ultrafiltration, or column chromatography using affinity labeled support. All depends.

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