Immunoglobulin quantitation and enumeration of immunoglobulin-producing cells: comparison of two index of B-cell activation.

Human lymphocytes were cultured under different conditions to determine the effects of technical variations on the response to pokeweed mitogen (PWM) and Staphylococcus aureus as measured by (a) quantitation of immunoglobulins (Ig) in the culture supernatants and (b) enumeration of Ig-secreting cells (ISC) by a reverse plaque technique. The highest numbers of ISC were measured when the cells were cultivated in standing tubes, without glutamine supplementation during the culture, and at a concentration of 1 x 10(6) cells/ml. The highest Ig concentrations were measured under similar conditions, except that somewhat higher values were obtained with cell cultivated at 1 x 10(6)/ml in 2 ml cultures with PWM stimulation and 2.5 x 10(6)/ml in 1 ml cultures with S. aureus stimulation. In time-course studies, peak ISC responses occurred on day 5 with each mitogen, whereas extracellular Ig levels kept rising until day 7, possibly owing to accumulation of secreted Ig. Measurements of the numbers of ISC and of secreted Ig levels in simultaneous cultures of lymphocytes from the same donors showed no correlation; however, co-stimulation of cultures with PWM and concanavalin A (to stimulate suppressor cells) depressed both Ig levels and ISC numbers. These results suggest heterogeneity in the plaque-forming cell population with respect to rate of Ig secretion, but indicate that these two techniques both reflect B-cell activation. They should not, however, be considered interchangeable, and the two probably should be used in conjunction for complete characterization of B-cell activation.