Saying I only have the protein purification gel report for 3 different preps with total soluble flow through and fractions 1-5 eluted with imidazole so is there any possible way I can tell whether the presence of more bands in the elutions are because of insufficient washing or truncated/degraded protein still having the his tag? The first prep was from a 3 hour induction at 30 c with 0.5 mM iptg and washed with 25 column volume 20 mM imidazole and 25 with 40 mM imidazole the second and third prep were washed with just 50 column volume of 20 mM imidazole and 40 was omitted cause the first prep showed that the 40 washed little amount of the desired protein. the culturing conditions were an O/N induction at 18C with 0.5 mM IPTG for the second and 0.1 mM IPTG 3 hour induction at 30 for the third. And I am trying to figure out whether the 40 mM imidazole is needed to wash non specifically bounded proteins as the second and third had more bands in the elutions.
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