Methods for the purification and quantitation of IgD

Abstract

Methods of isolation of IgD from D-myeloma and normal sera have been explored as has a sensitive method to quantitate low levels of IgD. DEAE-cellulose chromatography of a 30-45% saturated ammonium sulfate fraction of D-myeloma serum, yielded an IgD rich fraction in which IgG contamination did not exceed 20-25%. This was fractionated on G-200 Sephadex and IgD was eluted free of IgG in the ascending half of the first protein peak. Specific antisera to IgD were insolubilized by copolymerization and by coupling to an insoluble carrier and were used to specifically adsorb IgD from normal sera. IgD was eluted from the immunoadsorbant with solutions of high molarity. Other fractions containing IgG and IgD were purified by removing IgG vnth specific anti-IgG immunoadsorbants. IgD levels of less than 0.1 mg/ml were quantitated by a radioimmunosorbent method which depends on the competition between IgD in an unknown and 125-labelled IgD in binding to an insolubilized antiserum to IgD. Levels in excess of 0.1 mg/ml were quantitated in Cudin tubes