Reduction and s-carboxymethylation of proteins: microscale method

INTRODUCTIONReduction and alkylation of microscale amounts of protein (10-50 μg) require special attention. Classical methods for desalting and dialysis should be avoided due to large sample losses. Adding a small quantity of detergent to the protein solution during all manipulations can also reduce protein loss. This protocol describes a procedure for reduction and S-carboxymethylation of low-microgram amounts of protein. Samples are reduced and alkylated in small volumes (<200 μl) in the presence of detergent and desalted (and concentrated) by RP-HPLC in yields of >90%.