Analysis of B Cell Activation Requirements with TNP-Conjugated Polyacrylamide Beads

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Abstract
TNP-polyacrylamide beads of varying epitope density were used as antigens to stimulate normal adult spleen cells, neonatal spleen cells, and spleen cells from mice with the CBA/N immune defect. Lymphocytes from immune defective (CBA/N × DBA/2)F1 male mice or normal 2- to 3-week-old (C57BL/6 × DBA/2) F1 mice were unable to respond to TNP-polyacrylamide beads of low epitope density but responded quite well to high density TNP-polyacrylamide beads. Furthermore, immune defective F1 male spleen cells were shown to be unresponsive to TNP-Dextran yet mounted significant responses to TNP-Sephadex, an insoluble analog of TNP-Dextran. Use of an antiserum with anti-Lyb7.1 activity, which recognizes a mature subset of B cells, also pointed to qualitative differences in the stimulatory activities of highly substituted TNP-polyacrylamide beads and lightly substituted TNP-beads. This antiserum was able to inhibit responses to the low epitope TNP-polyacrylamide beads, which presumably activate a mature subset of B cells, but did not block responses to the highly conjugated TNP-beads, which appear to activate both immature B cells and mature B cells. In the course of these studies it became clear that TNP-polyacrylamide bead is not a truly thymus-independent (TI) antigen. Although it is able to stimulate significant responses in nude spleen cells, it is an ineffective immunogen when cultured with cells extensively depleted of T cells.

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