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ibed previously. Transduced cells were further sorted by fluorescence-activated cell sorting based on GFP positivity and termed parental 786-O RCC cells. To establish organ-derived cell lines, we injected parental 786O cells intracardially into SCID mice. Briefly, parental 786-O cells Flow Cytometry Cells were harvested by trypsin digestion and 26106 cells in PBS was incubated with anti-Cad11 anti
formed. After optimizing the digestion conditions, the majority of the PDIb'a'-hGCSF protein was cleaved by TEV protease. A second HisTrap HP column was then used to remove the PDIb'a' tag, undigested PDIb'a'-hGCSF, and TEV protease, which also contained a His6-tag. Cleaved hGCSF weakly bound to the Ni column and was eluted by 50 mM imidazole. An SDS-PAGE analysis revealed the absence of any conta
the hGCSF fusion protein at 30uC, whereas the solubilities of the Trx-, GST-, and His6-tagged proteins were low at this temperature. MBP is thought to act as a general molecular chaperone by binding to hydrophobic residues present on protein surfaces. MBP-tagged proteins can be easily purified with commercially available MBP-binding columns. PDI forms and breaks disulfide bonds of proteins in the