19 February 2021 6 7K Report

Hi there,

 I am trying to purify a recombinant His-tag protein from insect cell (baculovirus expression system) I used buffer containing %10 glycerol, 50mM SODIUM PHOSPHATE, 300mM NaCl , 20mM imidazole PH:8 , protein PI is 9. and went through sonication (10 sec on/1min off for 10cycles ). Sample was centrifuged at 16000rpm and finally filtered with 0.45 Micron. I used his trap column for purification (washing buffer 50mM SODIUM PHOSPHATE, 300mM NaCl , 20mM imidazole PH:8 elution buffer (50mM SODIUM PHOSPHATE, 300mM NaCl , 500mM imidazole PH:8 . Protein was dialyzed with amicon ultra 15 (100 kda). The size of protein is 234 kda. However, there is a bunch of unspecific bands along with the protein of interest even after dialysis.

Can anyone who has successfully purify recombinant protein from insect cell give me suggestions as to the best way to get rid of unspecific bands?

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