Anti-hapten Antibody Response Research Articles

Reaginic antibody formation in the mouse: VI. Suppression of IgE and IgG antibody responses to ovalbumin following the administration of high dose urea-denatured antigen

The major antigenic determinants in ovalbumin molecules (OA) were lost following denaturation in 8 M urea. The urea-denatured antigen (UD-OA) failed to combine with anti-OA antibody, but was capable of priming mouse T cells specific for OA. BDF 1 mice primed with alum-precipitated OA were given three intravenous injections of 100 μg UD-OA at 3, 5, and 7 days after the primary immunization. The treatment with UD-OA suppressed both IgE and IgG antibody responses to OA. The same treatment of OA-primed animals with intravenous injections of OA resulted in suppression of IgE antibody response but enhanced IgG antibody response. Intravenous injections of either OA or UD-OA suppressed both IgE and IgG anti-DNP antibody responses of DNP-OA-primed animals but failed to suppress anti-hapten antibody responses to DNP-keyhole limpet hemocyanin. The effect of the treatment on helper T cells and B memory cells in OA-primed animals was studied by adoptive transfer experiments. The results showed that the OA-treatment as well as UD-OA-treatment suppressed the development of both helper function of T cells and B memory cells in the spleen, but the UD-OA treatment was more effective than OA-treatment for the suppression of B cell development. Possible mechanisms for the suppression of the development of immunocompetent cells were discussed.

Induction of anti-hapten antibody response by hapten-isologous carrier conjugate: II. Specificity of hapten-reactive helper T cells

Anti-hapten antibody production was elicited by the immunization of hapten-isologous carrier conjugate (PAB-MGG) in mice. Spleen and lymph node cells taken from these primed mice could demonstrate their helper activity for anti-DNP antibody production when transferred intravenously into 600R X-irradiated recipient mice along with DNP-primed B cells and the double hapten conjugated carrier, DNP-MGG-PAB. Isologous carrier (MGG)-primed cells could not demonstrate this helper activity. Accordingly, helper cells reactive for a haptenic group are considered to develop by the immunization of hapten-isologous carrier conjugate. Hapten-reactive helper activity was also induced by the immunization of other hapten-isologous carrier conjugates, e.g., MAB-MGG, PABS-MGG or PAB-MSA. These hapten-reactive helper cells were T lymphocytes, as the helper activity of PAB-MGG-primed cells was completely abolished by in vivo ATS-treatment. Helper activity of PAB-MGG-primed cells for DNP-primed B cells was also demonstrated through the double hapten conjugated heterologous carrier DNP-HGG-PAB to be the same as with DNP-MGG-PAB, but weakly through DNP-KLH-PAB. As HGG but not KLH resembles MGG in composition, almost all hapten-reactive helper T cells can be considered to recognize not only haptenic groups but also physicochemical properties of the hapten-conjugated carrier site. However, these helper T cells could discriminate structural differences among related haptenic groups, because PAB-MGG-primed cells clearly responded to DNP-MGG-PAB to demonstrate their helper activity for DNP-primed B cells, but responded only weakly to DNP-MGG-PABS or DNP-MGG-MAB. When the specificity restrictions of T and B cells to the same haptenic group were compared by responsiveness measured after the antigenic stimulation (B cell function by anti-hapten antibody production and T cell function by helper activity), differences were noted, as PAB-MGG-primed T cells could respond not only to DNP-MGG-PAB but also fairly well to DNP-MGG-MAB to demonstrate their helper activity, but PAB-MGG-primed B cells responded to only PAB-MGG. Thus, hapten specificity appears to be much more restricted for B cells than T cells. The difference of this responsivity between B cells and helper T cells was thought to derive from the specificity difference of B cell and helper T cell receptors rather than from any sensitivity differences of the experimental procedure. The differences in the specificity restrictions of receptors of B and helper T cells were discussed in the light of hapten-specificity.

Effect of Nippostrongylus brasiliensis infection on anti-hapten IgE antibody response in the mouse : II. Mechanism of potentiation of the IgE antibody response to a heterologous hapten-carrier conjugate

Nippostrongylus brasiliensis infection was capable of potentiating the primary and secondary mouse IgE antibody response to a hapten coupled to a nonhelminthic antigen (ovalbumin, Ov). When mice were infected 14 or 5 days before an ip injection of 1 μ g dinitrophenylated Ov (DNP-Ov) mixed with 1 mg Al(OH) 3 , the anti-DNP IgE antibody response was greatly enhanced. The infection did not enhance anti-DNP IgG 1 antibody response. Potentiation of the primary IgE antibody response coincided with the generation of Ov or N. brasiliensis antigen (Nb) specific lymphoid cells of thymic origin (T cells). Moreover, it was found in adoptive transfer experiments that the potentiated response against DNP determinants can be obtained by concomitant administration of Nb with DNP-Ov into recipients that have been given the combination of lymphoid cells from infected and DNP-primed donors. These results suggest that Nb specific T cells could provide a certain signal, probably a soluble factor, upon stimulation with the worm antigen.

Cellular Cooperation During in Vivo Anti-Hapten Antibody Responses

Abstract Cellular interactions in adoptive secondary anti-hapten antibody responses to the hapten 2,4-dinitrophenyl (DNP) have been studied. It was shown that DNP-specific B cells must interact with carrier specific helper T cells to give optimal responses. Independent titration of B cell and helper cell activity in adoptive anti-DNP antibody responses gave the following results: Doubling the number of transferred B cells approximately doubled the subsequent antibody response. Doubling the number of helper cells leads to nearly 4 times as much anti-DNP antibody, measured 7 days after boosting (“premium effect”). This marked effect of helper cell number on the antibody response is thought to be due primarily to the interaction of two populations of carrier-specific cells in the helper effect, or to the interaction of two activities of a single population of helper cells, namely clone activation and clone expansion. Only a very small proportion of the premium effect given by helper cells could be attributed to increases in antibody affinity.

Cellular Cooperation During in Vivo Anti-Hapten Antibody Responses

Abstract The effect of x-irradiation on T and B cells involved in an in vivo adoptive anti-hapten antibody response has been studied. In vitro x-irradiation inactivates both B cells and T cells. B cells are somewhat more sensitive than T cells to such treatment, the dose required to reduce B cell activity by 50% being about 50 R, whereas 200 R will reduce helper cell activity by 50%. By contrast, if recipient mice are primed with carrier, they develop helper activity that is totally resistant to 500 R x-irradiation within 3 days of immunization. This activity can not be transferred from irradiated, carrier-primed mice by either spleen cells or serum, although anti-carrier antibody is shown to give a modest helper effect late after boosting. However, this radioresistant helper effect can be reproduced in a cell transfer system by giving carrier-primed spleen cells to normal recipients that are then x-irradiated (500 R) and given a large dose of carrier protein. Neither cells nor carrier alone will give rise to such radioresistant helper effects. Similar helper activity is occasionally observed when normal mice are irradiated 24 hr after being given helper cells, in the absence of carrier, and boosted shortly thereafter. The differences between these radioresistant helper effects, and the radiosensitivity of helper cells transferred after x-irradiation, may be related to x-ray induced changes in cell migration patterns.

Adjuvant activity of mycobacterial fractions. III. Adjuvant effect of cell wall of Mycobacterium bovis BCG on cell-mediated cytotoxicity in mice.

A quantitative assay and characterization of oil-attached cell wall of Mycobacterium bovis BCG (BCG-CWS) which stimulates cell-mediated immunity of spleen cells to alloantigens in mice were carried out by an in vitro cell-mediated cytotoxicity test using 51Cr-labeled target cells. C57BL/6J mice (H-2b) were immunized intraperitoneally with mastocytoma cells (H-2d) with or without oil-attached BCG-CWS. The cytotoxicity, comparable to that of spleen cells from mice immunized with mastocytoma cells (3 X 10(7)), could be induuced in spleens of mice immunized with a mixture of mastocytoma cells (10(4)) and oil-attached BCG-CWS. The enhancing effect persisted from 55 days or more after the alloantigenic immunization. Oil-attached BCG-CWS enhanced cell-mediated cytotoxicity of T cells in the spleen and the mesenteric lymph node, but not in the thymus. The cytotoxicity showed specificity toward the alloantigen used for immunization. In addition to BCG-CWS, the cell walls of Nocardia rubra and Corynebacterium diphtheriae PW8 and the peptidoglycolipids of Mycobacterium tuberculosis Aoyama B were found to be potent stimulants of cell-mediated cytotoxicity in mice. Oil-attached BCG-CWS did not enhance humoral response to mastocytoma cells but enhanced cell-mediated cytotoxicity when viable mastocytoma cells were used as antigen. The above result was supported by the fact that anti-hapten antibody response induced by viable trinitrophenyl (TNP)-mastocytoma cells (10(4)) plus oil-attached BCG-CWS did not increase to the maximum levels as was observed in mice immunized with a larger number of mastocytoma cells (3 X 10(7)) alone, while cell-mediated cytotoxicity induced by the same treatment increased to the maximum level obtained by immunization with mastocytoma cells (3 X 10(7)) alone.

Cellular Cooperation During in Vivo Anti-Hapten Antibody Responses

Abstract An adoptive secondary anti-2,4-dinitrophenyl (DNP) antibody response involving T-B cell collaboration has been studied. In particular, attempts have been made to affect the unexpectedly steep log dose-response curve obtained when graded numbers of helper cells are transferred to irradiated recipients given a fixed number of B cells (premium effect). A variety of means were used to alter helper cell activity, and this activity was then measured quantitatively, as was the ability of the helper cells present after these treatments to give a premium effect. It was shown that activated T cells are approximately twice as active as spleen cells in helper activity and give a comparable premium effect. Graded doses of anti-θ serum plus complement markedly reduce the helper activity of spleen cells without affecting the premium effect given by the residual cells. Treatment of primed cell donors with limited doses of heterologous anti-mouse thymocyte serum (ATS) before transfer does not affect B cell activity, but readily inactivates helper cells, again without affecting the premium effect given by the residual cells. Adult thymectomy (ATx) of helper cell donors before priming with carrier led initially to increased helper activity relative to agematched control donors. This increase may reflect the loss of nonspecific suppressor T cells from spleens shortly after ATx. Late after ATx, there was about a 2-fold decrease in helper activity, probably reflecting a loss of helper cell precursors. At no time was there any change in the premium effect. In view of the failure of any of the techniques used to abolish the premium effect given by helper cells in this response, it seems likely that this premium effect is due to the cooperative interaction of two very similar types of mature T cell. Alternatively, the premium effect observed here may result from the interaction of two activities of a single type of T cell which is mediated by independent factors.

Immunologic and Physicochemical Properties of Enhancing Soluble Factors for IgG and IgE Antibody Responses

Rabbits were immunized with dinitrophenyl-coupled Ascaris antigen (DNP-Asc) or ragweed antigen (DNP-Rag) included in aluminum hydroxide gel and their mesenteric lymph node cells were cultured for 24 hr in vitro in the presence of free homologous carrier. The cell-free supernatant thus obtained enhanced both IgG and IgE antihapten antibody responses of DNP-primed cells to DNP-heterologous carrier conjugate (DNP-keyhole limpet hemocyanin). Since the cell-free supernatant obtained from Rag-specific cells enhanced antibody response of hapten-primed cells raised by immunization with DNP-Asc, no carrier specificity was involved in the enhancement. It was found that treatment of primed cells with 10-5 M pactamycin suppressed the formation of the enhancing soluble factor, whereas the factor was readily formed in the presence of 2 mug/mol of cytosine arabinoside in the culture. The results indicated that cell proliferation was not required but de novo synthesis of protein was essential for the formation of soluble factor(s). The enhancing factor was not absorbed by either carrier-coated or anti-carrier antibody-coated immunosorbent. It was also found that the enhancing factor was formed by incubating primed cells with carrier-coated Sepharose. The cell-free supernatant containing no free carrier enhanced both IgG and IgE anti-hapten antibody responses. The activities of the cell-free supernatant to enhance IgG and IgE antibody responses were not absorbed by anti-Fab, anti-gamma-or anti-mu-chain antibody immunosorbent, indicating that the nonspecific enhancing factor did not possess immunoglobulin determinant. The cell-free supernatant was fractionated by sucrose density gradient ultracentrifugation and by gel filtration with three radiolabeled proteins, i.e., IgG, ovalbumin, and cytochrome C as markers. Enhancing activity for IgG antibody response was recovered in a fraction between ovalbumin peak (40,000 m.w.) and cytochrome C peak (20,000 m.w.). The activity for IgE antibody response was recovered in a fraction containing IgG marker (150,000 m.w.). By block electrophoresis, both activities were detected in beta globulin fraction. The results suggested that different T cell factors are involved in the IgG and IgE antibody responses.

Regulation of Antibody Response in Vitro

Abstract Attempts were made to induce antibody response of hapten-primed cells by stimulation with anti-immunoglobulin (Ig) antibody and cellfree supernatant (CFS) which contained nonspecific enhancing factor. Mesenteric lymph node cells were obtained from rabbits which were primed with dinitrophenylated ragweed antigen (DNP-Rag), and the primed cells were incubated in vitro with either anti-rabbit γ-chain or anti-Fab antibody for 24 hr. After washing, the cells were cultured in CFS which was obtained from the culture of DNP-Ascaris (DNP-Asc)-primed lymph node cells with free carrier (Asc). The results showed that hapten-primed cells were triggered for IgG anti-DNP antibody response by the two reagents, i.e., anti-Ig antibody and CFS, both of which did not share any antigenic specificity with the priming antigen (DNP-Rag). Neither anti-Ig alone nor CFS alone induced anti-hapten antibody response. Anti-Ig and CFS triggered not only hapten-primed B cells but also the other IgG-bearing (B) cells for IgG synthesis. Lymph node cells of unprimed animals were activated for IgG synthesis by the stimulation with anti-Ig followed by culture in CFS. Evidence was obtained that anti-Ig antibody has to be divalent for the activation of B cells. Stimulation of DNP-primed cells with the F(ab′)2 fragment of the antibody and CFS-induced anti-hapten antibody response, whereas the Fab′ fragment of the anti-Ig plus CFS failed to do so. The results suggested that bridging of cell-surface Ig receptors by multivalent ligand is the initial step of B cell activation. Another evidence for the activation of hapten-primed B cells by anti-Ig was obtained by supplemental immunization of DNP-Asc-primed animals with the Fc fragment of goat IgG. Stimulation of mesenteric lymph node cells from these animals with goat anti-rabbit-Ig alone (no CFS) resulted in the formation of anti-DNP IgG antibody. The results indicated that lymphocytes primed for goat IgG collaborated with DNP-primed IgG-B cells when the lymph node cells were stimulated with goat anti-Ig.

Effect of Nippostrongylus brasiliensis infection on anti-hapten IgE antibody response in the mouse : I. Induction of carrier specific helper cells

Inoculation of infective larvae of Nippostrongylus brasiliensis into A/J, BALB/c, and SJL mice primed intraperitoneally (ip) 3 weeks before infection with 1 μg of dinitrophenylated keyhole limpet hemocyanin (DNP-KLH) mixed with 1 mg Al(OH) 3 induced a carrier effect on anti-DNP IgE and IgG 1 antibody responses when the experimental mice were secondarily immunized with an ip injection of 1 μg of DNP-coupled N. brasiliensis extract (DNP-Nb) plus alum 2 weeks after infection. The magnitude of the hapten specific antibody response did not correlate rigidly with the number of larvae in the inoculum. Thus, a dose of 100 larvae was as effective in inducing the carrier effect as a dose of 800 larvae. Kinetic studies in A/J and BALB/c mice revealed that the anti-DNP IgE antibody response reached a maximum titer 7 days after the secondary immunization. These studies also showed that the enhanced IgE antibody response persisted for more than 40 days, while the response in all control groups terminated prior to that time. Using the adoptive transfer system, it was demonstrated that lymphoid cells obtained from the spleens or the mesenteric lymph nodes of infected mice cooperated with DNP-KLH primed cells to produce hapten specific IgE and IgG, antibodies when the challenge was made with DNP-Nb but not when it was made with 1 μg DNP-ovalbumin, clearly indicating carrier specificity. The helper activity of the cells obtained from infected mice was completely abolished or greatly reduced by the in vitro treatment with anti-θ serum and complement. The helper cells with maximum activity were present as early as 14 days after inoculation. The level of helper activity gradually decreased after 14 days. The results indicate that N. brasiliensis infection is effective in inducing carrier specific helper cells of thymic origin (T cells) in anti-DNP antibody responses. These results confirm those obtained by other investigators and add the new observation that N. brasiliensis infection elicits special helper T cells which induce an enhancement as well as a prolongation of anti-DNP IgE antibody response.

The immunological properties of haptens coupled to thymus‐independent carrier molecules. III. The role of the immunogenicity and mitogenicity of the carrier in the induction of primary IgM anti‐hapten responses

Hapten (DNP-lys) conjugates of two putatively nonimmunogenic polymers, hyalutonic acid and poly-gamma-D-glutamic acid, induce significant primary IgM anti-DNP responses in C3H mice. Preparations of various immunogenic (Type 3 pneumococcal polysaccharide (SIII), levan, E. coli lipopolysaccharide) and nonimmunogenic (hyaluronic acid and poly-glutamic acid) polymers were tested for their ability to act as polyclonal mitogens in vitro. In serum-containing spleen cell cultures, only lipopolysaccharide stimulated substantial cell proliferation. In serum-free medium, and using high specific activity [3H]thymidine, lipopolysaccharide, levan, SIII and to a lesser degree hyaluronic acid induced significant thymidine incorporation. However, under the latter conditions cell survival and proliferation were much less impressive. There was no apparent correlation between the capacity of various polymers to induce lymphocyte proliferation and their "potency" as carriers for the generation of a primary IgM anti-DNP response. Furthermore while low doses of lipopolysaccharide elicited "polyclonal" antibody formation in vivo, high doses of SIII, levan and hyaluronic acid did not. These results indicate that T cell-independent B cell triggering is dependent on the polymeric nature of the antigen, and that polymers need not be immunogenic or mitogenic to act as carriers for the induction of primary IgM anti-hapten antibody responses.

Regulation of Homocytotropic Antibody Formation in the Rat

Abstract An antigen-specific subcellular component that augments anti-hapten IgE class homocytotropic antibody (HTA) response was extracted from thymocytes and spleen cells of rats hyperimmunized with dinitrophenylated Ascaris suum extract (DNP-As) or with Ascaris protein (As) alone. This soluble component, when injected into neonatally thymectomized rats, restored the ability to produce anti-DNP HTA and hemagglutinating antibodies of the recipients that were otherwise incapable of producing antibodies upon immunization with DNP-As. The thymocyte extract from either normal rats or those hyperimmunized with DNP-bovine serum albumin (DNP-BSA) did not show the restoring ability for both HTA and hemagglutinating antibodies. Furthermore, this “helper” activity was largely absorbed by the carrier protein, As, but not with hapten, DNP-BSA, indicating the helper component possesses specificity and affinity for the carrier molecule. Although these antigen-specific characteristics of the helper component are much like those of the previously reported “inhibitory” component of thymus-derived lymphocytes that specifically suppresses the ongoing HTA response, a number of differences in their immunochemical and physicochemical properties were observed. Unlike the inhibitory component, the helper component possessed Fab and µ-chain determinants as revealed by absorption studies. The molecular weight of the helper component was between 200,000 and 100,000 daltons, the value much larger than that obtained for the inhibitory component. These results indicate that IgT-like molecule is involved in the induction of in vivo HTA response, and further suggest that HTA formation is regulated by two distinct antigen-specific subcellular components of thymus-derived lymphocytes in the rat.

Dissociation of T Cell Helper Function and Delayed Hypersensitivity

Abstract The effect of tolerization with varying doses of deaggregated protein carrier on two T cell functions—delayed hypersensitivity (DHS) to the carrier and helper functions for the secondary anti-hapten antibody response to a hapten carrier conjugate—was studied. It was found that high doses of tolerogen inhibit the development of both DHS and helper function, while low doses of tolerogen inhibit the development of helper function but not of DHS.

Induction of anti-hapten antibody response by hapten-isologous carrier conjugate: I. Development of hapten-reactive helper cells by hapten-isologous carrier

Anti-hapten antibody production was elicited by the immunization of hapten-isologous carrier conjugate (DNP-MγG) in mice. The spleen and lymph node cells taken from those primed mice were effectively stimulated with hapten-heterologous carrier conjugates (DNP-KLH and DNP-BαA) as well as hapten-homologous carrier conjugate (DNP-MγG) when transferred into X-irradiated recipient mice. The reactivity of DNP-MγG-primed cells to DNP-heterologous carrier conjugates was not due to the mutual crossreactivity of the carrier with MγG on cellular level, since the spleen and lymph node cells primed with DNP-KLH or DNP-BαA could only be stimulated with corresponding hapten-homologous carrier conjugate. The responsiveness of DNP-MγG-primed cells to hapten-heterologous carrier conjugates was due to the result that hapten-reactive helper cells were developed by the immunization of hapten-isologous carrier and these cells cooperated with hapten-specific B cells. The helper activity of the hapten-isologous carrier-primed cells was resistant to 600-R X-irradiation in vitro and sensitive to in vivo ATS treatment. This suggests that the helper activity induced by hapten-isologous carrier is of T cell origin. The helper activity of hapten-isologous carrier-primed cells was also developed by the immunization of PAB-MγG, and clear cooperative interaction between PAB-MγG-primed cells and DNP-specific B cells was demonstrated through DNP-MγG-PAB. The possible mechanism of helper cell development induced by the immunization of hapten-isologous carrier conjugate was discussed in light of the hapten specificity of helper activity.

Hapten-Specific IgE Antibody Responses in Mice

Abstract The studies presented in this paper were performed to compare the responses of B lymphocytes of the IgG and IgE classes, respectively, to relatively constant conditions of activated T cell participation in antibody production by these cells. Two systems were employed: a) regulation of antibody responses by a potent nonantigen-specific T cell influence provided by the reaction of allogeneic T cells with alloantigens on the surface of antigen-specific B cells (“allogeneic effect”); and b) induction of adoptive secondary cooperative anti-hapten antibody responses by administering hapten and carrier determinants on separate molecules. The data presented here demonstrate that the allogeneic effect, although providing a potent mechanism for stimulating IgG B lymphocytes, either fails to stimulate or suppresses IgE B cells under the same conditions. Furthermore, under circumstances where antibody responses are elicited by administering nonlinked hapten and carrier determinants, the evidence implies that IgE B cells can be stimulated to a greater extent than IgG B cells in conditions of limited T cell participation. Taken collectively, these data support the concept that differences observed in the degree of helper function for one or the other antibody class may reflect differences in sensitivities of IgE vs IgG B cells to the complex enhancing and suppressive regulatory influences of the same T cell populations rather than to the existence of different T cells for different immunoglobulin classes.

Reaginic Antibody Formation in the Mouse

Abstract An adoptive transfer system for an IgE anti-hapten antibody response was established in DBA/1 mice with dinitrophenylated ovalbumin (DNP-OA) and keyhole lympet hemocyanin (DNP-KLH) as antigens. Maximal anti-DNP IgE antibody response was obtained by transfer of DNP-primed cells and carrier-specific cells for the secondary carrier to irradiated syngeneic mice, followed by challenge with DNP-secondary carrier conjugate in saline, a procedure which does not induce a primary IgE antibody response. Recipients of hapten-primed cells or carrier-specific cells alone showed meager or no IgE antibody response. Effect of aluminum hydroxide gel (alum) with antigen for carrier immunization and hapten priming was studied by the adoptive transfer technique. It was found that immunization of donors of carrier-specific cells with OA in saline, which did not induce either IgG or IgE antibody response, resulted in a high level of helper function for both IgE and IgG anti-hapten antibody responses in the adoptive response of DNP-KLH-primed cells to DNP-OA. Similarly, addition of a low dose alum with KLH was sufficient for carrier priming for helper function with respect to the anti-DNP IgE antibody response. It was shown, however, that alum had a significant effect on hapten priming. Recipients of the spleen cells from donors primed with DNP-OA or DNP-KLH plus a high dose (1 mg) alum gave much higher anti-hapten IgE antibody response than the recipients of DNP-primed cells obtained by priming with the same protein in saline. Animals primed with DNP-KLH with a low (50 µg) dose alum gave IgG but no IgE secondary response. In spite of the presence of both carrier-specific helper cells and hapten-primed precursors for IgE antibody response, these animals gave rise to a secondary IgG antibody response but failed to show IgE antibody response upon a booster injection with DNP-KLH in saline. If their spleen cells were transferred to irradiated animals, the recipients showed high IgE and low IgG antibody responses upon injection of DNP-KLH in saline. These results suggest that secondary IgE antibody response is enhanced in irradiated recipients.

Regulation of Antibody Response in Vitro

Abstract Mesenteric lymph node cells were obtained from rabbits that were primed with dinitrophenylated ragweed antigen (DNP-Rag) and supplemented with free Ascaris antigen (Asc); in vitro anti-hapten antibody response of the primed cells was studied. It was found that stimulation of Rag-sensitized cells with free Rag enhanced anti-DNP antibody response to DNP-unrelated carrier conjugate (DNP-KLH) but suppressed the response of the same cell suspension to DNP-Asc. It was also found that both IgG and IgE antibody responses to DNP-Rag were suppressed by the addition of free Asc with the homologous antigen. Mechanisms involved in the enhancement and suppression of antibody response by free carrier were explored with cell-free supernatant (CFS) obtained from the culture of primed lymph node cells with free carrier. The results showed that essentially all CFS preparations studied enhanced antibody response when they were added to primed lymph node cells which had been treated with a hapten-protein conjugate. Some of the CFS preparations, however, suppressed antibody response of the same primed cells, if they were added to the cells together with the antigen. Evidence was obtained which indicated that suppression of the antibody response by CFS is not due to an excess amount of enhancing soluble factor. Several preparations of CFS were incubated with the same primed cell suspension, either before or after incubation with antigen, to determine their suppressive effect and enhancing effect. The results showed that suppressive effect of CFS on the unstimulated cells did not correlate with their enhancing effect on the antigen-stimulated cells. The experiment eliminated the possibility that the same T cell factor expresses two opposite effects depending on the stage of B cell differentiation, and suggested the presence of both suppressive and enhancing soluble factors in the preparations of cell free supernatant. The experiment also showed that enhancing soluble factor for IgE antibody response was obtained from carrier-specific cells primed with alum-precipitated antigen but the cells proliferated by the immunization with the carrier included in complete Freund's adjuvant did not release the enhancing soluble factor for IgE. It was also found that enhancing soluble factor for IgG antibody neither enhanced nor suppressed IgE antibody response.

Enhancing factor on anti-hapten antibody response released from PPDs-stimulated tubercle bacilli-sensitized cells

There is a considerable body of evidence that cellular interaction is required for the induction of antibody response to various antigens (Claman and Chaperon, 1969); Miller and Mitchell, 1969; Taylor, 1969). It is now well established in hapten-carrier system that a good anti-hapten antibody response is obtained when hapten-committed antibody forming cell precursor (AFCP) cooperates with carrier-committed helper cells through hapten-carrier conjugate. The former cells are derived from bone marrow and have the immunoglobulin-like receptor specific for the haptenic group on their cell surface. The latter cells are thymus-derived and have a receptor specific for the carrier moleculr and help anti-hapten AFCP to produce anti-hapten antibody. We reported previously that tubercle bacilli-sensitized cells have helper activity on the secondary anti-hapten antibody formation in the presence of hapten-conjugated PPDs in vivo culture system (Takatsu et al., 1972 a). The helper activity of tubercle bacilli-sensitized cells were diminished when the cells were treated with θ-antibody and guinea-pig complement prior to cell transfer (Takatsu et al., 1972 b). It is however, still obscure how both cells of the tubercle bacilli-sensitized cells and hapten-specific AFCP interact to produce anti-hapten antibody in the presence of hapten-tuberculin protein conjugate. In the present study, we found that a soluble factor is released from tubercle bacilli-sensitized cells in vivo and in vitro, and enhances effectively anti-hapten antibody response of a hapten-carrier-primed cells in the presence of a heterologous carrier conjugated with the hapten.

Enhancing factor on anti-hapten antibody response released from PPDs-stimulated tubercle bacilli-sensitized cells

Enhancing factor on anti-hapten antibody response released from PPDs-stimulated tubercle bacilli-sensitized cells

Hapten-Specific IgM and IgG Antibody Responses in Mice against a Thymus-Independent Antigen (DNP-Salmonella)

2,4-Dinitrophenyl hapten, when coupled to <i>Salmonella choleraenuis</i> bacilli (DNP-Salm), was capable of inducing sequential IgM and IgG antihapten antibody responses in the mouse, both of which were found to be totally thymus-independent. The antigen could also elicit a secondary antihapten antibody response of both IgM and IgG classes in mice preimmunized with the hapten coupled to either homologous or heterologous carrier. No ‘helper’ or ‘suppressor’ acitivity was found in salmonella-primed T cells. However, this antigen was not capable of inducing IgE antibody response even with the aid of adjuvants. Furthermore, the time-dependent increase in antibody affinity (maturation) did not occur following repeated injections of DNP-Salm.