Immunocyte Response to a Purified Bacterial Toxin (Choleragen) and Toxoid: Cytokinetics, Immunoglobulin Class, and Specificity

Abstract

Abstract A unique indirect plaque-forming cell (PFC) assay has been developed to assess the immune response to choleragen, a highly purified exo-enterotoxin derived from Vibrio cholerae cultures, at the cellular level. Spleen cells releasing anti-choleragen antibody were readily detectable with choleragen-sensitized sheep erythrocytes as “target” cells in a variation of the localized hemolysin in agar technique. Injection of 1-µg doses of the toxin into BALB/c mice was sufficient to stimulate large numbers of specific antibody PFC. Heat-treated choleragen preparations, i.e., procholeragenoid and choleragenoid, also induced a detectable immune response at 1-µg doses; however, the results indicate that the PFC response varied as a function of the dose and form of the cholera antigen used for immunization. Choleragen and procholeragenoid induced essentially similar immune responses and were more immunogenic than choleragenoid, although all immunogens stimulated IgM (19S) and IgG (7S) PFC. Peak IgM PFC responses were detected on days 6 and 7 whereas peak IgG PFC responses occurred 1 day later in all cases. A booster injection of choleragen (1 µg) increased the numbers of IgM and IgG PFC approximately 4- to 5-fold. IgA PFC were also detected after a booster injection. Specificity experiments revealed that the toxin and toxoid, but not a vibrio whole cell vaccine or lipopolysaccharide preparation, could inhibit the anti-choleragen PFC response.