Refolding single-chain antibody (scFv) using lauroyl-l-glutamate as a solubilization detergent and arginine as a refolding additive

Abstract: Therapeutic potential of immunoconjugates has opened a new window for antibody-based biopharmaceuticals. Greater tissue penetration and hence enhanced cell toxicity are obtained with a smaller version of antibodies. While the whole antibody can be readily produced via mammalian expression system, antibody fragments often require refolding of insoluble proteins. Here we report a new refolding method for antibody fragments using a novel amino acid–based detergent as a solubilizing agent and arginine-assisted refolding. Inclusion bodies of antibody fragments were solubilized by 2.5% lauroyl-l-Glu (C12-l-Glu) and successfully refolded by multi-step dilution into a buffer solution containing arginine hydrochloride and thiol/disulfide-exchange reagents. Adjustment of temperature was found to be critical for increase in the refolding yield. Although each protein requires appropriate optimization, solubilization by C12-l-Glu and dilution refolding assisted by arginine can generate the native, functional antibody fragments. The procedure should enable us to utilize bacterial expression systems for the large-scale manufacturing. [Copyright &y& Elsevier]