Antibody production in vitro. The effects of anti-allotypic serum on the secondary response of rabbit spleen cell suspensions to sheep erythrocytes (SRC).

Rabbit anti-allotypic sera directed against rabbit immunoglobulins stimulated an increase in the number of direct PFCs in suspensions of cells from the spleens of rabbits killed after a boost of SRC. This effect occurred within the first 24 hours of incubation of these cells with antiserum in the absence of antigen in the culture medium. High levels of these antisera were much less stimulatory than low during the early period of culture, but more prolonged incubation with antiserum in high concentrations resulted in increased numbers of PFC in these cultures. Amounts of antisera which were stimulatory in the absence of SRC in the culture medium were found to inhibit the stimulatory effect of antigen in PFC during the first 1–2 days of culture. Like high concentrations of anti-allotypic sera alone, this early inhibition was followed by a stimulatory increase of PFC on more prolonged culture in the presence of both SRC and anti-allotypic serum. Anti-allotypic sera affected the levels of PFC in these studies even in the absence of any change in the rate of DNA synthesis, as measured by the incorporation of [H]thymidine. Further to this, it was found that concentrations of antisera which stimulated increased uptake of [H]thymidine during 24–48 hours of culture, actually depressed the levels of PFC in these cultures. It was, therefore, concluded that the action of anti-allotypic sera on PFC in these cultures was by means of a direct effect on the process of production of antibodies by these cells and was not dependent on the stimulation of cell division in the antibody-producing population. This stimulatory effect of anti-allotypic sera was prevented by Actinomycin-D and the inhibitory effects of complement on PFC in cultures incubated with anti-allotypic sera indicated that antibodies had combined with receptor molecules on these cells. The effects of the combination of SRC and anti-allotypic sera on the levels of PFC would suggest that the same cell population in these cultures was being affected by both these agents and that the receptor molecule for them was specific antibody. The effect of anti-allotypic sera of the PFC in these rabbit spleen cell suspensions was shown to have some degree of non-specificity. Since normal rabbit serum as well as rabbit antiserum against a OX19 did not have any effect it was considered possible that this non-specific stimulation of PFCs was also due to antibodies directed against antigenic determinants in rabbit immunoglobulins.