Factors affecting the immunoperoxidase demonstration of intracellular immunoglobulins and J chain from cytocentrifuged cell smears.

Abstract

Immunohistochemical methods were used to study 1) the optimum fixation conditions for the preservation of human J chain and immunoglobulin (Ig) immunoreactivity and 2) the relation of J chain synthesis by plasmablasts and plasma cells to Ig synthesis in cell smears of cultured human peripheral blood lymphocytes stimulated with pokeweed mitogen (PWM). J chain was demonstrated using the indirect immunoperoxidase method, and intracellular Ig was demonstrated with the unlabeled antibody--enzyme method. In the sequential double staining procedure, J chain was demonstrated using the indirect immunoperoxidase method followed by the demonstration of Ig with the direct immunofluorescence method. Optimum preservation of J chain immunoreactivity was obtained with fixation in neutral buffered formalin at 22 degrees C for 5 min followed by immediate immunoperoxidase staining. False negative results were seen when the slides were stained 2 weeks after fixation. In PWM-stimulated smears, J chain appeared on day three, simultaneously with or after the onset of Ig synthesis. In double stained smears most IgG-positive cells also showed immunoreactivity for J chain from the third day on.