The failure to show a necessary role for C3 in the in vitro antibody response

Abstract

The in vitro antibody response of mouse spleen cells to TNP coupled to both T‐dependent and T‐independent carriers as well as to sheep erythrocytes has been studied to investigate the possible role of complement activation in the induction of antibody formation. The following has been found. (1) In vitro responses of both IgM and IgG can be obtained to both T‐dependent and T‐independent antigens in serum‐free media, although they are smaller than those found in serum‐containing media. This shows that no exogenous source of complement is necessary for in vitro antibody formation by spleen cells. (2) Similarly, normal antibody responses are obtained if the cultures are grown in human serum depleted of C3b‐inactivator, which contains high concentrations of C3b. (3) In the presence of antibody to mouse C3 the response to the T‐independent antigen is reduced, the IgM responses being more affected than the IgG. However, purified F(ab′)₂ anti‐C3 has no inhibitory effect and it therefore seems likely that it is the formation of intact immune complexes containing Fc rather than the interference with C3 function that is responsible for the inhibition seen. (4) The conventionally purified anti‐complementary factor from cobra venom has no effect on the antibody response in serum‐free culture or when human or fetal calf sera are used. In no experiment was any potentiation of T‐dependent responses observed. However, the presence of quite small concentrations (2 %) of cobra venom factor (CVF)‐treated normal mouse serum produced a profound inhibition of the antibody responses, affecting IgG responses more than IgM, and being T‐dependent more than T‐independent, but no part of the response being unaffected. However, CVF heated to 70 °C which destroys the anticomplementary activity but not the contaminating phospholipase A was equally effective. More highly purified CVF, on the other hand, was not. These findings strongly suggest that the inhibition observed is due to residual phospholipase A reacting with lecithin in the mouse serum rather than to any anticomplementary effect. (5) Spleen cells taken from mice treated with CVF in vivo were also unable to give a full IgG response to the T‐dependent antigens if challenged 24 h after CVF treatment. However, after 48 h no inhibition was observed. It therefore seems to us that it is unlikely that C3 plays any real part in the in vitro antibody response. Our data – like the occurrence of genetically C3‐deficient humans with apparently normal antibody responses – would seem to preclude any necessary role for C3 in antibody production.