Characteristics of immunoglobulin secretion in man evaluated by a reverse hemolytic plaque assay

The kinetics of Pokeweed mitogen-induced transformation of human B-lymphocytes into immunoglobulin-secreting cells were examined in a reverse hemolytic plaque assay using protein A-coated sheep red blood cells as indicator cells. Peak responses occurred consistently after 6-8 days of incubation. After determination of the optimal experimental conditions the RHPA was found to be a reliable tool to estimate ISC in vitro; the technique was also found to be applicable for experiments surveying the B-cell response of an individual over a period of months. The PWM-induced transformation of B cells was absolutely T-cell-dependent. Other substances known as typical T-cell mitogens were also tested for polyclonal B-cell activation and some of them showed significant responses. Further experiments have shown that co-cultivation of non-HLA identical cells did not increase the number of plaques in unstimulated cultures whereas the addition of PWM leads to the generation of ISC within the expected range. These findings open a wide field of application of the RHPA in experimental and clinical immunology.