T lymphocyte-mediated suppression of myeloma function in vitro. II. Evidence for regulation of hapten-binding myelomas by syngeneic hapten- specific cytolytic T lymphocytes

BALB/c splenocytes stimulated in vitro with trinitrophenyl (TNP)-modified syngeneic cells inhibit the secretion of antibody by the TNP-binding BALB/c myeloma MOPC 315 in the presence of soluble TNP-Keyhole limpet hemocyanin (KLH). The effector cells are hapten-specific, H-2-restricted, Thy-1.2-bearing, Ly-2-positive T lymphocytes whose precursors are resistant to pretreatment with cyclophosphamide. These phenotypic properties are typical of hapten-specific cytolytic T lymphocytes (CTL). The TNP-reactive CTL that inhibit MOPC 315 cells fail to suppress H-2d myelomas that do not bear TNP-specific surface receptors, and this is not attributable to differences in total binding of TNP-KLH to the different myeloma cells. Moreover, azobenzene arsonate (ABA)-specific CTL inhibit MOPC 315 cells in the presence of the double conjugate TNP-ABA-KLH, but not in the presence of soluble TNP-KLH or ABA-KLH. These results show that H-2-restricted, hapten-specific lymphocytes regulate the function of myeloma cells that bind the hapten only to specific surface receptors, and provide a model for associative recognition of surface H-2 determinants and receptor-bound antigen. The results are discussed with reference to the mechanisms of T lymphocyte-target cell interactions, and the possible physiologic role of hapten-reactive CTL in specifically regulating anti-hapten antibody responses.     

Recognition of polymorphic H-2 domains by T lymphocytes. I. Functional role of different H-2 domains for the generation of alloreactive cytotoxic T lymphocytes and determination of precursor frequencies

In the present communication, the repertoire of alloreactive cytotoxic T lymphocytes (CTL) clones was quantitatively investigated by limiting dilution analysis and by target inhibition with a panel of monoclonal antibodies (mAb). These mAb have previously been shown to define two distinct alloantigenic domains, A and B, on the H-2Kk molecule. The Poisson distribution analysis of H-2Kk-specific CTL clones generated in a limiting dilution system revealed three CTL populations with different precursor frequencies. The high frequent population is suppressed by an unknown suppressive mechanism that allows less frequent CTL populations to become visible. Target inhibition studies with a panel of Kk-specific mAb showed that these CTL populations differ not only in their precursor frequency but also in their specificity for different H-2 epitopes on the Kk molecule. Thus clones of the high frequency population are almost exclusively specific for determinants within domain A. In contrast, the low frequency population displays predominant specificity for determinants of domain B, while the population with medium frequency is blocked equally well by mAb against either domains A or B. Each mAb blocked only a fraction of clones indicating that each CTL subpopulation may consist of a large number of clonotypes with specificity for different H-2 epitopes. The data suggest that CTL recognize basically the same polymorphic domains on the H-2Kk molecule defined by antibodies, and they show that regulatory mechanisms determine the expressed repertoire in CTL populations.     

Idiotypes on major histocompatibility complex-restricted virus-immune cytotoxic T lymphocytes

Specificity-associated determinants could be demonstrated on major histocompatibility complex (MHC)-restricted virus-immune cytotoxic T lymphocytes (CTL) using antisera raised in syngeneic mice. This result indicates that the number of clonotypes sufficiently small to allow the detection of such idiotypic determinants. The functional properties of three anti-idiotypic antisera were tested in blocking studies and by idiotypic antisera plus complement. Whereas the former test did not reveal any results obtained from binding studies, i.e., all three anti- idiotypic antisera specifically reacted with CTL of the type used for immunization and had no effect on syngeneic influenza-immune CTL, syngeneic alloreactive CTL, or NDV-immune CTL of an H-2-disparate strain. These results are discussed in terms of current models for MHC- restricted CTL responses.     

Antigen recognition by H-2-restricted cytolytic T lymphocytes is not mediated by two independent receptors

Detergent-solubilized murine cytolytic T lymphocytes (CTL) clones were incorporated into Sendai virus-containing synthetic liposomes. When these liposomes were then fused with other CTL clones possessing a different non-cross-reacting specificity, the fusion products were observed to lyse target cells recognized by both parental CTL clones. This method was then used with two H-2-restricted CTL clones of different, non-cross-reacting specificities (anti-H-2b-H-Y or anti-H-2b Moloney leukemia virus). Once again, the fusion products were found to be lytic against both target cells recognized by the parental clones, but in no instance was there any observable lysis of target cells bearing the same nominal antigen in the context of different H-2 molecules. These results provide strong evidence that antigen recognition by H-2-restricted CTL is not mediated by two independent antigen receptors.     

Clonal heterogeneity in the functional requirement for Lyt-2/3 molecules on cytolytic T lymphocytes: analysis by antibody blocking and selective trypsinization

While it is well established that murine cytolytic T lymphocytes (CTL) express the Lyt-2/3 molecular complex on their surface, conflicting results have been reported concerning the role of this complex in CTL activity. In the present study this question was reinvestigated at the clonal level. Although different (H-2b anti-H-2d) CTL clones expressed comparable amounts of Lyt-2/3 molecules, as assessed by quantitative flow microfluorometry, the activity of some clones was inhibited by low doses (10 ng) of monoclonal anti-Lyt-2 or anti-Lyt-3 antibodies (in the absence of complement), whereas other clones were not inhibited by either antibody at doses as high as 5 microgram. Treatment of these clones with doses of trypsin sufficient to cleave Lyt-2/3 antigenic determinants from the cell surface resulted in a similar dissociation: clones that were inhibited by antibodies lost cytolytic activity, whereas "uninhibited" clones were unaffected by trypsin treatment. Moreover, the dissociation observed among different alloreactive clones could be demonstrated with self-H-2-restricted (H-2b anti-MSV) clones exhibiting cross-reactivity with normal H-2d products. The lytic activity of these clones against the relevant syngeneic target cells was unaffected by anti-Lyt-2 antibodies or trypsin, whereas their cross- reactivity on H-2d target cells was abolished by either treatment. These results provide direct evidence for clonal heterogeneity in the requirement for Lyt-2/3 molecules in CTL-mediated lysis. It is proposed that the function of Lyt-2/3 molecules is to stabilize the interaction between CTL receptors and the corresponding antigens on the target cells and that the requirement for such a stabilization is correlated with low number and/or affinity of CTL receptors.     

Allorestricted cytotoxic T cells. Large numbers of allo-H-2Kb- restricted antihapten and antiviral cytotoxic T cell populations clonally develop in vitro from murine splenic precursor T cells

Cytotoxic T lymphocyte (CTL) responses of splenic T cells from C57BL/6 B6) mice and mutant H-2Kbm1 (bm1) mice to haptenic (trinitrophenyl [TNP] ) and herpes simplex virus (HSV) determinants in the context of an allogenic (wild-type or mutant) H-2Kb molecule were analyzed in a modified limiting dilution system. In the B6-anti-bm1TNP mixed leukocyte reaction (MLR), estimated frequencies for precursors of CTL clones that lysed bm1TNP targets ranged from 1/120 to 1/400; in the bm1-anti-B6TNP MLR, estimated frequencies of precursors of CTL clones that lysed B6TNP targets ranged from 1/500 to 1/1,300. Estimated frequencies for precursors of CTL clones that lysed the respective unmodified and TNP-modified allogeneic targets were two- to three-fold lower. Lytic specificity patterns determined by split-well analysis showed that at least 20-30% of the generated CTL populations (selected for a high probability of clonality) in both MLR displayed allorestricted lysis of TNP-modified concanavalin A blast targets. In the B6-anti-bm1HSV MLR, estimated frequencies for precursors of CTL clones that lysed bm1HSV targets ranged from 1/70 to 1/300; in the bm1-anti-B6HSV MLR, estimated frequencies for precursors of CTL clones that lysed B6HSV targets ranged from 1/300 to 1/1,200. Again, estimated frequencies for precursors of CTL clones that lysed the respective noninfected and virus-infected allogeneic targets were two- to fourfold lower. Of the CTL populations selected for a high probability of clonality at least 30-60% displayed allorestricted lysis of virus-infected lipopolysaccharide blast targets in both MLR. It is concluded that a large fraction of clonally developing CTL populations stimulated with TNP-modified or HSV-infected allo-H-2Kb-bearing cells displayed an allorestricted pattern of recognition. It was further evident that the estimated frequencies of splenic precursors that generated allorestricted CTL clones was two- to threefold higher than the estimated frequencies of precursors that gave rise to the respective alloreactive CTL populations.     

Allospecific cytolytic T lymphocytes recognize conformational determinants on hybrid mouse transplantation antigens

Alloreactive cytolytic T cell (CTL) lines and clones have been used to identify the sites of polymorphism of antigens of the major histocompatibility complex (MHC). Specific CTL were generated against wild-type H-2b products by cells from H-2b mutant mice that had one or a few amino acid changes in either the alpha 1 or alpha 2 domains of the Kb or Db class I molecules. These CTL populations, which might be expected to react with determinants expressed on single MHC domains, were examined for lytic activity on L cells expressing newly constructed hybrid class I molecules. Transformed cell lines expressing native class I molecules or hybrid class I molecules in which the alpha 1 and alpha 2 domains of H-2Kb had been substituted by those domains of H-2Db were lysed by H-2Db-specific CTL. Similarly, all H-2Kb-specific CTL recognized hybrid molecules in which the alpha 1 and alpha 2 domains of H-2Kb were inserted into the H-2Db molecule. In contrast, exchange of the alpha 1 domains of H-2Kb and H-2Db resulted in a total loss of recognition by Kb and Db-specific CTL. These results suggest that the allodeterminants recognized by H-2 mutant CTL are influenced by interactions between the alpha 1 and alpha 2 domains, findings similar to those seen using conventional alloreactive T cells (11). These results were compared to the binding of alloreactive mAbs, including 5 new mAbs specific for the Kb molecules. Finally, it was shown that primary and secondary CTL responses could be generated by direct sensitization against hybrid class I molecules, demonstrating that these molecules express neoantigenic determinants recognized by alloreactive CTL.     

Specificity studies on cytolytic T lymphocytes directed against murine leukemia virus-induced tumors. Analysis of monoclonal cytolytic T lymphocytes

The specificities of cloned cytolytic T lymphocytes (CTL) were studied for the analysis of CTL populations generated against murine leukemia viruses (MuLV) in H-2 congenic BALB/c (H-2d) and BALB.B (H-2b) mice. In particular, CTL generated in response to tumors induced by Gross MuLV and Friend MuLV were studied; these tumors expressed virus-induced antigens that do not cross-react and that can be distinguished from each other. The systematic study of 92 CTL clones clearly indicated that MuLV-immune CTL were highly heterogeneous with respect to both the intensities of target cell lysis that they mediated and to their specificity of recognition of MuLV-induced tumor target cells. Various categories of CTL clones were identified, ranging from CTL clones tht were tightly H-2 restricted and specific for the immunizing tumor to CTL clones that displayed no discernible patterns of specificity and that attacked a large number of different target cells. In addition, the surface markers of these cloned CTL were defined, and the best conditions for their prolonged maintenance in culture were determined. The present data indicate that future efforts in the definition of target antigens recognized by tumor-specific CTL should be performed with monoclonal lymphocytes.     

Simultaneous expression of H-2-restricted and alloreactive recognition by a cloned line of influenza virus-specific cytotoxic T lymphocytes

Based on theoretical considerations and several types of experimental evidence with heterogeneous cell populations it has been proposed that alloreactive T cells and major histocompatibility complex (MHC)-restricted T cells directed to foreign non-NHC antigens represent overlapping subsets. In this report we provide direct evidence for this hypothesis at the clonal level. We have isolated a cloned continuous influenza virus-specific cytotoxic T cell (CTL) line derived from a single (H-2b X H-2d)F1 CTL precursor which simultaneously exhibits H-2-restricted cytotoxicity of influenza A/Japan/305/57 virus in association with H-2Kd and alloreactive cytotoxicity for H-2Kk alloantigen. Cold target inhibition data demonstrate that both MHC-restricted and alloreactive recognition is mediated by the same cell population. In addition to cross-reactivity at the target cell level, we shown that this cloned CTL line can be specifically stimulated to proliferate either by A/Japan/305/57 virus-infected cells expressing H-2Kd or by uninfected cells of the H-2Kk haplotype.     

Fine specificity of alloimmune cytotoxic T lymphocytes directed against H-2K. A study with Kb mutants

The fine specificity of alloimmune cytotoxic T lymphocytes (CTL) was investigated in CTL responses across the smallest known H-2 differences, those based on mutation at a single H-2 locus. CTL were generated in all possible mixed lymphocyte culture (MLC) combinations among seven H-2Kb mutants and the mouse strain of origin, C57BL/6 (B6-H-2b). CTL were also generated in all F1 hybrid responder/homozygous stimulator-cell combinations among four Kb mutants and B6-H-2b. CTL activity was measured in cell-mediated lympholysis (CML) against target cells from all Kb mutants and B6-H-2b. Cross-reactivity against targets other than the MLC stimulator was extensive and led to the distinction of 64 CML target determinants. Two Kb mutants (B6-H-2bm6 and B6.C-H-2bm9) showed identical typing for all 64 CML determinants. CML reactions after MLC between these two haplotypes were mutually negative. The mutants B6-H-2bm3 and B6.C-H-2bm11 showed identical typing for 47 of the 64 determinants. Their close relationship is in agreement with the finding that H-2bm3 anti-H-2bm11 CTL were the only ones that exclusively lysed target cells of stimulator-cell genotype. On the basis of CML typing, the sequence of relatedness of the mutants with H-2b is as follows: bm6/bm9-bm10-bm3-bm1-bm11, bm6/bm9 being the closest to, and bm11 the most distant from H-2b. The extensive cross-reactivity of alloimmune CTL appears to reflect immunogenetic complexity rather than lack of specificity. Comparison with other reports supports the notion that the system of Kb CML determinants, recognized by alloimmune CTL, is at least partially overlapping with the H-2Kb specificity repertoire involved in the associative T cell recognition of virus-infected cells.     

T lymphocyte-mediated suppression of myeloma function in vitro. III. Regulation of antibody production in hybrid myeloma cells by T lymphocytes

To investigate the mechanisms by which T lymphocytes regulate myeloma function in vitro, the effects of regulatory T cells on antibody secretion by a hybrid myeloma cell line were examined. Suppressor T cells (Ts) specific for idiotypic determinants on M315 (IgA, lambda 2 anti-2,4-dinitrophenol and anti-2,4,6-trinitrophenol [TNP]) and MPC 11 (IgG2b, kappa) myeloma proteins inhibit antibody secretion by the appropriate parental myeloma cells. When cocultured with a hybrid cell line derived by fusion of MOPC 315 and MPC 11 myelomas, the idiotype-reactive Ts inhibit secretion of only the immunoglobulin (Ig) bearing the relevant idiotype. In contrast, syngeneic TNP-reactive cytolytic T lymphocytes (CTL) inhibit antibody secretion by TNP-binding MOPC 315 cells but not by MPC 11 cells in the presence of soluble TNP-keyhole limpet hemocyanin (KLH), and this inhibition probably represents a prelytic effect of the CTL. Such TNP-reactive CTL, in the presence of TNP-KLH, inhibit both IgA and IgG secretion by the MOPC 315-MPC 11 hybrid, which is consistent with a prelytic effect. Thus, myeloma hybrids are a useful tool for investigating the effector function of regulatory T cells. These results are discussed with reference to the mechanisms of action of regulatory T cells and their relevance to modulation of physiologic humoral immune responses.     

Specific adsorption of IgM antibody onto H-2-activated mouse T lymphocytes

Evidence is presented to support the contention that IgM demonstrable by surface immunofluorescent staining on H-2-activated T cells represents specifically adsorbed B-cell-derived alloantibody. T cells activated to H-2 determinants expressed surface IgM only when the progenitor cell populations contained B lymphocytes. IgM was not detected on T cells activated to determinants which fail to stimulate alloantibody production (e.g., M-locus determinants). In addition, IgM-negative H-2 activated T cells (derived from B-cell-depleted lymphoid cells), unlike M-locus-activated T cells, adsorbed IgM in a specific manner when incubated in vitro with "early bleed" antisera raised against the activating H-2 determinants.     

Cytotoxic T lymphocytes induced against allogeneic I-region determinants react with Ia molecules on trinitrophenyl-conjugated syngeneic target cells

The major histocompatibility complex codes for determinants which are recognized by and serve as targets for cytolytic T lymphocytes (CTL) (1). Antigens coded for by the K and D loci of the H-2 complex can activate xenogeneic or allogeneic CTL (2,3). In addition, the H-2K or H-2D gene products function as those molecules against which syngeneic CTL responses specific for chemical, viral, and minor H antigens are directed (4-8). It has recently been shown that Ia determinants can also serve as target antigens for distinct but weaker CTL responses (9-13). Those clones which recognize Ia antigens see them independently of K- or D- coded antigens as shown in genetic studies and by antisera-blocking experiments (12,13). We have proposed that the existence of clones of CTL specific for I-region-coded determinants is not fortuitous; rather these clones specifically recognize Ia determinants and may have an immunoregulatory role. These CTL may affect those immune functions which are at least partially dependent on or controlled by I-region-coded molecules. Two predictions can be made and tested concerning the role of Ia determinants in cytolytic systems and the role, if any, of I-region- specific CTL in regulating the immune response: (a) that if as we and others have shown, certain Ia specificities can serve as a third series of major histocompatibility antigens, then Ia antigens should be susceptible to the same types of antigenic modifications as H-2K- or H-2D-coded structures and thus serve as targets for CTL directed against modified-self in selected systems; and (b) that allogeneically induced I-region-specific CTL should demonstrate cross-reactivity with targets bearing modified syngeneic I-region-coded determinants. Data will be present which demonstrates that trinitrophenyl (TNP)-modified syngeneic I-region determinants can serve as targets for CTL induced by allogeneic Ia antigens.     

Cytotoxic T lymphocyte responses in allogeneic radiation bone marrow chimeras. The chimeric host strictly dictates the self-repertoire of Ia- restricted T cells but not H-2K/D-restricted T cells

The present report has used fully H-2 allogeneic radiation bone marrow chimeras to assess the role of host restriction elements in determining the self-specificity of Ia- and H-2K/D-restricted T cells that participate in the generation of trinitrophenyl (TNP)-specific cytotoxic T lymphocytes (CTL). It was demonstrated that there exists a stringent requirement for the recognition of host thymic-type Ia determinants, but there exists only a preference for host thymic-type H-2K/D determinants. Indeed, once the stringent requirement for recognition of host Ia determinants was fulfilled, anti-TNP CTL were generated in response to TNP-modified stimulators that expressed either donor-type or host-type H-2K/D determinants. The CTL that were generated in response to TNP-modified donor-type stimulators were shown to be specific for TNP and restricted to the non-thymic H-2K/D determinants of the chimeric donor. Thus, these results demonstrate in a single immune response that the thymic hypothesis accurately predicts the self-specificity expressed by Ia-restricted T cells, but does not fully account for the self-specificity expressed by H-2K/D-restricted T cells. These results are consistent with the concept that H-2K/D-restricted T cells, but not Ia-restricted T cells, can differentiate into functional competence either intrathymically or extra-thymically. The present results are also informative for understanding the cellular interactions that are required for the generation of antigen-specific CTL responses. The Ia-restricted T cells that are required for the generation of H-2K/D-restricted anti-TNP CTL were shown to be helper T (TH) cells since (a) like TH cells functioning in antibody responses, they were specific for Ia determinants expressed by accessory cells, and (b) their function could be replaced by either TNP-primed, irradiated TH cells or by nonspecific soluble helper factors. It was also shown that the T-T cell interaction between Ia-restricted TH cells and H-2K/D-restricted precursor CTL (pCTL) is not Ia restricted. Rather, the results demonstrate that the generation of anti-TNP CTL responses involve two parallel sets of major histocompatibility complex-restricted cell interactions, an Ia-restricted TH-accessory cell interaction required for TH cell activation, and an H-2K/D-restricted pCTL-stimulator cell interaction required for pCTL stimulation. The interaction between activated TH cells and stimulated pCTL is mediated, at least in part, by nonspecific soluble helper factors.     

The induction of virus-specific cytotoxic T lymphocytes with solubilized viral and membrane proteins

Reconstituted membranes were prepared from detergent solubilized P815 (H-2d) tumor cell membranes and solubilized Sendai virus protein. These reconstituted membranes stimulated a virus-specific H-2-restricted secondary CTL response. Stimulating activity was dependent upon the presence of both viral and P815 protein in the same membrane and was restricted to the H-2 specificity present in the reconstituted membrane. Liposomes prepared from solubilized Sendai virus proteins and partially purified H-2 alloantigen also had activity for CTL induction. The results demonstrate the feasibility of using detergent solubilized membrane proteins to study antigen recognition by virus-specific, H-2 restricted cytolytic T lymphocytes.     

Separate cytotoxic T lymphocyte subsets recognize the different H-2 specificities

Using a monolayer adsorption technique, the fine specificity of cytotoxic effector T lymphocytes (CTL) generated against allogeneic or semi-allogeneic H-2 haplotypes was investigated. The results show that: (a) CTL reacting with the private specificity expressed on an H-2.K molecule can be separated from those reacting with the public specificities expressed on the same molecule and (b) the CTL that recognize cross-reacting H-2 determinants (public specificities) can also be separated into several subpopulations. These data support the hypothesis that an allogeneic stimulation induces a large number of independent T cell clones that react with H-2 determinants.     

Antigen recognition by cloned cytotoxic T lymphocytes follows rules predicted by the altered-self hypothesis

Radiation chimeras prepared by injecting H-2 heterozygous F1 stem cells into lethally irradiated parental hosts show a marked, but not absolute, preference for host-type H-2 antigens in the H-2-restricted cytotoxic T lymphocyte (CTL) response to minor histocompatibility (minor H) antigens. We have selected for the anti-minor HCTL that are restricted to the parental H-2 type absent from the chimeric host and found that in two out of eight cases, such CTL lysed target cells of either parental H-2 type. From one of these CTL populations that lysed H-2d and H-2k target cells expressing BALB minor H antigens, clones were derived and further analyzed. The results showed that: (a) lysis of both H-2d and H-2k target cells was H-2 restricted; (b) H-2d restriction mapped to Dd, and H-2k restriction mapped to Kk; (c) testing against various H-2d and H-2k strains of different and partially overlapping minor H backgrounds as well as against the appropriate F1 crosses revealed that in Dd- and Kk-restricted killing, different minor H antigens were recognized. In a second system, a CTL population was selected from normal (H-2d x H-2k)F1 mice that was specific for H-2d plus minor H antigens and for H-2k plus trinitrophenylated bovine serum albumin. We interpret these findings in terms of the altered-self hypothesis: The association of one H-2 antigen with one conventional antigen X may be recognized by the same T cell receptor specific for the complex formed by a different H-2 antigen in association with a second conventional antigen Y. The implications of these observations for the influence of self H-2 on the generation of the T cell receptor repertoire are discussed.     

Characterization of antigen-specific, Ia-restricted, L3T4+ cytolytic T lymphocytes and assessment of thymic influence on their self specificity

The goals of the present study were: (a) to generate antigen-specific L3T4+ cytolytic T lymphocytes (CTL), (b) to determine their major histocompatibility complex (MHC) restriction specificity, and (c) to assess the influence of thymic MHC determinants on their self specificity. We found that L3T4+ CTL specific for either trinitrophenyl (TNP)-modified self determinants or minor histocompatibility antigens could be generated from Lyt-2- responder T cells provided that the response cultures were supplemented with supernatants rich in helper factors. Such antigen-specific L3T4+ CTL were Ia-restricted by the criteria that they lysed only Ia+ target cells and that their lysis of Ia+ target cells was specifically inhibited by anti-Ia monoclonal antibodies. The relative frequency of L3T4+ pCTL was found to be only 5-10% of the total anti-TNP pCTL present in the spleens of normal mice. Finally, we utilized radiation bone marrow chimeras to assess the influence of the thymic haplotype on the self-Ia specificity of L3T4+ CTL. Both bulk culture and limiting dilution experiments revealed that the self-Ia specificity of L3T4+ anti-TNP CTL from F1----parent and A----B allogeneic chimeras was not markedly skewed toward the haplotype of the chimeric thymus. These results contrast with those obtained previously for L3T4+ anti-TNP Th cells and demonstrate that in the radiation bone marrow chimera model of T cell differentiation, the self specificity of Th cells but not pCTL is markedly influenced by the haplotype of the chimeric thymus.     

Alloantigen receptors on activated T cells in mice. I. Binding of alloantigens and anti-idiotypic antibodies to the same receptors

B6 alloantigens in supernates from one-way mixed lymphocyte reaction (MLR) cultures of AKR T cells against B6 lymph node cells rebound specifically to (B6)AKR-T-cell blasts after overnight incubation and recovery of these blasts from trypsin treatment. A similar specificity was observed with the binding of SJL alloantigens to (SJL)AKR-T-cell blasts. In both cases, the corresponding alloantigens were rebound several-fold more efficiently than control alloantigens. In a different assay system, T-cell receptors were studied with anti-idiotypic sera. These antisera were raised by repeated injection of purified (B6)AKR-T-cell blasts into (AKR X B6)F1 hybrid mice. These F1a(AKRaB6) sera reacted with the majority of (B6)AKR-T and (B6)(AKR X SJL)F1-T-cell blasts. They also reacted with a lower, though sizable, number of (SJL)AKR-T- and (SJL)(AKR X B6)F1-T-cell blasts. No reaction was observed with (B6)SJL-T, concanavalin A-activated AKR T-cell blasts, normal B6, (AKR X B6)F1, and (AKR X SJL)F1 T cells. Normal AKR T cells were positive only minimally above background. F1a(AKRaB6) sera could be made specific for (B6)AKR-T and (B6)(AKR X SJL)F1-T-cell blasts by absorption of contaminating antibodies with (SJL)AKR-T-cell blasts. Finally, it was shown by competition experiments that the receptors on MLR-activated T-cell blasts that bind alloantigens were the same as those binding anti-idiotypic antibodies. In addition, it was found that at least a fraction of alloantibodies share common idiotypic determinants with receptors on MLR-activated T-cell blasts.     

Alloantigen-specific idiotype-bearing receptors on mouse T lymphocytes. I. Specificity characterization and genetic association with the heavy- chain IgG allotype

The present study describes the qualitative reactions of a xenogeneic anti-idiotype (Id) antiserum produced in a mouse-gamma-globulin-tolerant rabbit (5,936) against B6 anti-CBA IgG antibodies. The results showed that such an anti-Id antiserum reacts specifically against anti-H-2k antibodies and against H-2k alloantigen-activated T cells from the following pairs of congenic mice: B10 (H-2b) and B10.D2 (H-2d); and A.BY (H-2b) and A.SW (H-2s), but not against C3H.SW (H-2b) and C3H.OH (H-2o); and BALB/b (H-2b) and BALB/c (H-2d). CB 20 (BALB/c mice with the Ig-1b allotype) anti-CBA T blasts also express idiotypic determinants that react with rabbit 5,936 antiserum. Thus, positive reactions are obtained between rabbit 5,936 anti-Id antiserum and anti-H-2k IgG preparations and T blasts from mice carrying the Ig-1b or Ig-1e allotype, but not from mice carrying the Ig-1a allotype. These reactions are qualitatively independent of the H-2 genotype of the Id-producing mice. Such a finding strongly suggests that the Id-bearing receptor molecules on mouse T cells are coded for by genes that are associated with the Ig heavy-chain-linkage group and not to the mouse histocompatibility complex. Furthermore, the anti-Id antibodies studied react preferentially against anti-H-2k antibodies or T cells with specificity toward the IAk-region-associated serological specificities. Thus, genes associated with the Ig heavy-chain-linkage group seem to be structural genes for at least T-cell receptors with specificity for IA-region-coded membrane antigens.