In vitro induction of a Haemophilus influenzae type b polysaccharide-specific antibody response in human peripheral blood lymphocytes of individuals recently vaccinated with an oligosaccharide-protein conjugate.

Abstract

In this paper an in vitro culture system for the induction of an antibody response to the Haemophilus influenzae type b polysaccharide (PRP) is described. Anti-PRP IgM and IgG antibody-secreting cells (ASC) and anti-diphtheria toxoid (DT) IgG ASC were detected in cultures of blood B and T cells derived from donors 4 to 6 wk after immunization with Haemophilus influenzae type b oligosaccharide-mutant diphtheria toxin (CRM197) conjugate (HbOC) and required in vitro restimulation with HbOC. When lymphocytes from HbOC-vaccinated donors were stimulated with PRP, anti-PRP IgM and IgG ASC could be detected in 50% offGe cases. Lymphocytes from PRP-vaccinated donors or non-vaccinated donors consistently failed to generate anti-PRP antibodies after in vitro stimulation with HbOC. Optimal in vitro responses were observed at concentrations of 0.06 to 0.6 micrograms/ml of Ag. At higher doses of Ag (6 micrograms/ml) anti-PRP and anti-DT antibody responses were suppressed. The in vitro generation of anti-PRP and anti-DT ASC, as detected by a spot-forming cell assay was shown to be T cell dependent, Ag dependent, and Ag specific. This culture system provides a model for the study of human B cell activation and immunoregulation by polysaccharide-protein conjugates and polysaccharides.