A new specific quantitation-in-gel method differentiating commercial human serum standards intended for RID analyses

A rapid, sensitive technique is described which measures small amounts of protein applied on agarose gel containing specific antibody. Alternating current through the gel reduces the influence of diffusion, and specific immunoprecipitate spots are formed. Their density after staining is proportional to applied protein. Quantitation of IgG and IgA in human serum standards was highly reproducible (P less than 0.001, Spearman coefficient of rank correlation test), and as little as about 35 ng Ig could be detected. Results are available within hours. Human sera and some purified preparations of 7S IgG and secretory IgA were grouped according to immuno-gel filtration findings. The group of samples containing fragments, aggregates, or unusually small amounts of monomeric IgG or IgA was also differentiated by specific immunoprecipitate spot (SIS) analysis from those mainly monomeric in character (Wilcoxon rank test, P less than 0.02). A World Health Organization reference serum (67/97) was used as standard. The study indicates that a human serum pool stored in aliquots at -20 degrees C is a good standard for quantitating serum IgG and IgA and that purified preparations are no better. It is suggested that the SIS assay could be advantageously applied to screening biologic fluids for unusual amounts or types of IgG and IgA.