Separation of immunoglobulin M (IgM) essentially free of IgG from serum for use in systems requiring assay of IgM-type antibodies without interference from rheumatoid factor

Abstract

The proposed method was designed to replace the tedious and difficult separation of immunoglobulin M (IgM) from IgG by sucrose gradient sedimentation. In this method, a 250-microliter portion of serum diluted 20-fold was passed through a small column of quaternary aminoethyl-Sephadex A-50 ion exchanger. IgG was not retained, but additional washes were required to remove all but 5%. A second buffer-eluting fluid recovered an average of 80% of the original IgM in a defined dilution. The entire operation took 15 min. The efficiency of this process was evaluated by the following: (i) radial immunodiffusion measurements of IgG and IgM; (ii) recovery studies of isohemagglutinins; and (iii) demonstrated removal of interference by the rheumatoid factor. The method was applied successfully to distinguish rubella IgM antibody.