Crossed immunoelectrophoresis and electroimmunoassay of human IgG subclasses

Abstract

Human IgGl, IgG2, IgG3 and IgG4 in WHO pool 67/97, a normal serum pool, Cohn Fraction II and individual sera were examined by crossed immunoelectrophoresis and electroimmunoassay in agarose with IgG subclass specific rabbit antisera. In these methods the fact that IgG subclasses differ in the electrophoretic field is utilized: IgG4 is located anodically, IgG3 cathodically, and IgG2 and IgG1 both anodically and cathodically. The mean, S.D. and range of serum IgG1, IgG2, IgG3 and IgG4 in 20 normal adults found by the electroimmunoassay were given and related to the amount in the WHO pool 67/97. The IgG subclasses values obtained by electroimmunoassay agreed with the values obtained by single radial diffusion. The reproducibility of double determinations (interplate variations) was 1.5--5.5 per cent. Repeated freezing, thawing and storage of the serum at room temperature did not influence quantitation of IgG subclasses. Cohn Fraction II was found to contain smaller amounts of IgG1, IgG2, and IgG4 than those found in the normal serum pools. Crossed immunoelectrophoresis and electroimmunoassay also easily reveal failing quality of IgG subclass antisera. To obtain good antisera in rabbits against IgG subclasses immunization should be done with several myeloma proteins with different electrophoretic mobility within the same subclass.