AbstractWe have isolated milligram quantities of active single chain antibody from the insoluble fraction ofEscherichia colicultures. The system relies on high‐level expression from a T7 RNA polymerase‐directed gene construct, 8 M urea to dissolve the desired protein out of the insoluble fraction, presumably inclusion bodies, isolation and concentration of the desired protein by nickel chelate IDA‐Ni(II) immobilized metal‐ion affinity chromatography (IMAC), and removal of urea from column fractions by dialysis directly into storage buffer. Routinely, about 50 of the protein loaded onto an IMAC column is recovered as single chain Fv at a concentration of approximately 0.7 mg/mL. As little as 3 days are required to obtain 10 mg of final product when starting with an overnight i
展开▼
