Preparation of Fab fragments from a mouse monoclonal IgM

A procedure is described for preparing and purifying Fab fragments from a mouse monoclonal IgM. The IgM was digested with trypsin in the presence of the reducing agent, cysteine. The resulting Fab fragments were alkylated with iodoacetamide and purified using a Sephacryl S-200 column. The Fab fragments had a molecular weight of 48 000 as determined by SDS polyacrylamide gel electrophoresis and molecular sieve chromatography. This procedure has been successfully used with five different mouse monoclonal IgM. The Fab fragments bound antigen with the same specificity as the whole IgM.