A U.S. Patent for “Tandem Folding Methods to Improve Protein Folding Yield” was awarded to Purdue University on April 28th, 2020. The present invention provides a method of refolding a solubilized cysteine-containing protein to its native form, the method comprising adding a reversible reducing agent to a solution containing the solubilized cysteine-containing protein to initiate folding of the cysteine-containing protein, wherein the reversible reducing agent is added at a ratio of [reversible reducing agent]:[thiols in the cysteine-containing protein] of 3:1 to 4:1; and adding an irreversible reducing agent to the solution at a time after the initiation of folding to breakdown aggregates of the cysteine-containing protein. Learn More