Double labeled-antigen method for demonstration of intracellular antigens in paraffin-embedded tissues

A double "labeled-antigen" method has been developed for the simultaneous staining of both kappa and lambda light chains in fixed paraffin sections. The method is a two step procedure utilizing a mixture of antisera against kappa and lambda light chains in the first stage, followed by the addition of a mixture of kappa antigen labeled with horseradish peroxidase and lambda antigen labeled with alkaline phosphatase. The selection of substrates yielding reaction products of contrasting color enabled the observer to distinguish kappa-containing cells (brown) from lambda-containing cells (blue). Reactive plasma cells stained either pure brown (kappa) or clear blue (lambda) in a ratio of 1.5:1. Blood vessels containing serum immunoglobulins showed a mixed brown-blue reaction, as did the Reed-Sternberg cells of some cases of Hodgkin's disease. The advantages of this double labeled-antigen method over previously reported methods for achieving double staining are discussed.