100 Î¼l supernatant from the supernatant was taken for the SDS-PAGE (ELUTION 1). The beads were resuspended in with 250 mM imidazole in a similar volume of buffer to that of the beads used. Spinning was done for 1 minute at 700g. All the supernatant was retained, and a further 100 Î¼l taken from the supernatant for SDS-PAGE (ELUTION 2). The beads were resuspended in the same volume of buffer with 1 M imidazole, and spinning accomplished for 5 minutes at 700g. All the supernatant was retained.
100 Î¼l from the supernatant for SDS-PAGE was taken and a similar volume of buffer added back to the pelleted beads. The contents were then gently agitated for 1 minute. Boiling was then accomplished for 5 minutes at 100oC, followed by 5 minutes of spinning at 700g. All the supernatant was retained. 100 Î¼l from the supernatant for SDS-PAGE was then taken and every sample ran on SDS page.
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