title: An improved haemolytic plaque assay for the detection of cells secreting antibody to bacterial antigens.
abstract: Recent advances in the development of conjugate polysaccharide vaccines for human use have stimulated interest in the use of assays detecting antibody-secreting cells (AbSC) with specificity for bacterial antigens. Here we present improved haemolytic plaque-forming cell (PFC) assays detecting AbSC with specificity for tetanus and diphtheria toxoid as well as for Haemophilus influenzae type b and pneumococcal capsular polysaccharides. These assays were found to be less time consuming, more economical and yielded 1.9-3.4-fold higher plaque numbers than traditional Jerne-type PFC assays. In the case of anti-polysaccharide AbSC of the IgG isotype, the increase was as high as 7.4-11.8 times. Evidence is presented that the pronounced improvement in the detection of the latter is due to the presence of aggregating anti-IgG antibody from the beginning of the assay. It is proposed that in the case of low affinity of anti-polysaccharide antibodies aggregation of secreted monomeric antibody (IgG) is critical for plaque formation and increases the avidity of binding to target cells.
authors.affiliation: Laboratory of Medical Immunology, Medical Department TTA, Rigshospitalet, Copenhagen, Denmark.
authors.forename: T
authors.initials: T
authors.lastname: Barington
authors.forename: C
authors.initials: C
authors.lastname: Heilmann
mesh_terms: D000906:Antibodies / Q000276:immunology; D000917:Antibody Formation; D000921:Antibody-Producing Cells / Q000276:immunology; D000942:Antigens, Bacterial / Q000276:immunology*; D006192:Haemophilus Infections / Q000175:diagnosis; D006193:Haemophilus influenzae; D006462:Hemolytic Plaque Technique; D006801:Humans; D011008:Pneumococcal Infections / Q000175:diagnosis; D012680:Sensitivity and Specificity; D013742:Tetanus / Q000175:diagnosis; D013745:Tetanus Toxoid
chemical_list: D000906:Antibodies; D000942:Antigens, Bacterial; D013745:Tetanus Toxoid