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Ni-NTA Superflow Columns - automated large-scale purification of 6xHis-tagged ,,, proteins

tallography. Currently, methods like FPLC are used for the purification of such amounts. The Ni-NTA Superflow Column procedure enables automated purification of up to 24 samples each containing up to 15 mg 6xHis-tagged protein (Table 1). Figures 1 and 2 show the high reproducibility and absence of cross-contamination in the automated Ni-NTA Superflow Column purification procedure. Ni-NTA Superflow Columns can also be used for manual gravity-flow purification of 6xHistagged proteins.

Cross-ContaminationFree Processing Highly Reproducible Yields Figure 1 SDS-PAGE analysis of twelve 6xHis-tagged protein preparations performed in parallel using Ni-NTA Superflow Columns. For E. coli culture volumes and yields see Table 1. CAT: 6xHis-tagged chloramphenicol acetyl-transferase; TNF- alpha: 6xHis-tagged tumor necrosis factor- alpha; 46: 6xHis-tagged 46 kDa protein; 38: 6xHistagged 38 kDa protein; GFP: 6xHis-tagged green fluorescent protein; 42: 6xHis-tagged 42 kDa protein. Aliquots (5 l) of the final eluates (3 ml) were loaded onto the gel. Proteins were visualized by Coomassie
staining. M: markers. Figure 2 SDS-PAGE analysis of twelve 6xHistagged green fluorescent protein preparations. Cleared lysates from 100 ml E. coli cultures were processed in parallel using Ni-NTA Superflow Columns. Aliquots (5 l) of the final eluates
(3 ml) were loaded onto the gel. Proteins were visualized by Coomassie staining. M: markers.

Table 1. Protein yields from preparati
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