Showing papers in "European Journal of Immunology in 1984"

The immunoglobulin heavy chain variable region (Igh‐V) locus in the mouse. I. One hundred Igh‐V genes comprise seven families of homologous genes

Abstract: Seven families of Igh-V genes have been defined by Southern blot analysis of genomic DNA from eighteen inbred strains of mice. Each of twenty-four cloned Vh genes hybridized to one of seven nonoverlapping sets of Eco RI restriction fragments. These families contain from 2 to approximately 40 hybridizing fragments. From these data we estimate that the mouse Igh-V locus consists of one hundred Vh genes. Genes within a Vh family share greater than 80% sequence homology while the sequence homology between families is generally less than 70%. There is extensive restriction fragment length polymorphism among the strains analyzed allowing the assignment of complete (Igh-V + Igh-C) Igh haplotypes for eighteen inbred mouse strains.

Cross-protection in mice infected with influenza A virus by the respiratory route is correlated with local IgA antibody rather than serum antibody or cytotoxic T cell reactivity.

Abstract: Mice previously infected with an aerosol of A/Rec 31 influenza virus were strongly protected against an aerosol challenge with A/Vic influenza as judged by lung virus titers recovered 2 days after the challenge infection. Such complete homotypic immunity was not achieved by priming with live Rec 31 virus injected i.v. or UV-inactivated Rec 31 virus administered s.c. together with Al(OH)3 and saponin. The reason for the superior protective effect of the natural infection was investigated. The protection induced by respiratory infection with Rec 31 virus was specific for influenza A viruses. It was not correlated with specific serum hemagglutination inhibition antibody titer or cross-reactive cytotoxic T (Tc) cell reactivity. Moreover, the transfer of splenic and lymphoid T cell populations with strong secondary Tc activity did not significantly reduce lung virus titers in recipient mice 3 days after infection. The protection however occurred in parallel with the presence of cross-reactive IgA antibody in the lung washings. It thus appears that local secretory IgA plays a causal role in the prevention of cross-infection by influenza A virus. Serum antibody and Tc cells, on the other hand, may be crucial for recovery from such infection. All mice primed with live Rec 31 virus, administered i.v. or by aerosol and expressing equally high levels of Tc reactivity, survived a lethal challenge with A/PR8 virus. The same challenge, however, killed half of the mice immunized s.c. with inactivated Rec 31 virus which induced only a low level of Tc reactivity.

MRC OX-19: a monoclonal antibody that labels rat T lymphocytes and augments in vitro proliferative responses.

Abstract: A mouse monoclonal antibody, designated MRC OX-19, has been prepared that binds to virtually all rat thymocytes, T lymphocytes and a maximum of 2% B lymphocytes. Immunoperoxidase studies on tissue sections showed no reactivity with nonlymphoid cells in intestine, thymus, spleen and lymph node. The antigen recognized by MRC OX-19 antibody was identified by metabolic and cell surface labeling of thymocytes followed by immunoprecipitation and sodium dodecyl sulfate gel elec-trophoresis. It is a surface glycoprotein of Mr = 69000. When included in in vitro assays for T cell functions, MRC OX-19 increased proliferation stimulated by allogeneic spleen cells or the lectins phytohemagglutinin and concanavalin A. The antibody itself was not mitogenic and its stimulatory effect could be correlated with an increase in interleukin 2 production. Taken together the data suggest that MRC OX-19 could be the equivalent of mouse Ly-1 antigen and human T1 antigen.

Protection of mice against the intracellular bacterium Listeria monocytogenes by recombinant immune interferon.

Abstract: Immune interferon, available at high specific activity through recombinant DNA technology, is known to activate macrophages to intra- and extracellular cytotoxicity. We now report that murine recombinant IFN-gamma activates macrophages to cytotoxicity also when applied in vivo. Furthermore, recombinant IFN-gamma can protect mice in vivo against the intracellular bacterial pathogen Listeria monocytogenes in a local as well as in a systemic infection model. The role of T lymphocyte-produced lymphokines in acquired resistance to facultative intracellular pathogens and their possible involvement in novel immunotherapy are discussed.

Monoclonal antibodies to stromal cell types of the mouse thymus

Abstract: Seven hybridoma cell lines secreting monoclonal antibodies (mAb) to nonlymphoid cells of the mouse thymus have been prepared. These mAb clearly demonstrate the heterogeneity of the thymic stroma. Based on their anatomical distribution patterns observed with the immunoperoxidase technique on frozen tissue sections, they were subdivided into four groups. The first group of mAb, ER-TR1, 2 and 3, detects antigens encoded for by the I region of the major histocompatibility complex. These antigens are expressed on both stromal and lymphoid cells in lymphoid organs. mAb of the second category, ER-TR4, react with epithelial cells in the thymic cortex. mAb of the third group detect stromal cells of the thymic medulla. One antibody of this group, ER-TR5, exclusively reacts with medullary epithelial cells. ER-TR6, the other antibody of this group, reacts with medullary interdigitating cells and macrophages. The fourth type of antibodies, ER-TR7, detects the reticular fibroblasts of the thymus. The possible role of the thymic cell types detected by the present antibodies in T cell differentiation is discussed.

Reactions among IgM antibodies derived from normal, neonatal mice.

Abstract: Attempting to obtain a representative sample of the "natural antibody" repertoire in the developing immune system, we have derived IgM-secreting hybridomas from 4 normal untreated BALB/c mice of the same litter on day 6 after birth. Partially purified IgM preparations obtained in the supernatants of 70 such clones were each screened in binding assays for reactivity with a panel of 9 IgM antibodies, randomly selected from the same collection. Five of these 9 IgM antibodies were found to react with a considerable number of other IgM in the collection, while the other 4 showed only sporadic reactivity. On the other hand, more than half of the 70 antibodies were found to bind specifically to at least one of these five. With a few exceptions, these reactions showed quantitative levels ranging from 5 to 20% of those observed between either of the two interacting IgM and monoclonal rat anti-mu antibodies. The selectivity of these reactions indicated V-region specificity, which was confirmed by analyzing in some detail the reaction between 2 IgM antibodies isolated from the same mouse.

Effect of cloned interferon-gamma on expression of H-2 and Ia antigens on cell lines of hemopoietic, lymphoid, epithelial, fibroblastic and neuronal origin.

Abstract: Interferon-gamma (IFN-gamma), tested in the form of the product of a cloned murine IFN-gamma gene, was found to increase the expression of H-2 antigens on cultured cell lines of a wide variety of cell types including factor-dependent hemopoietic cells, pre-B and B cells, macrophages, T cells, mast cells and cell lines of epithelial, fibroblastic and neuronal origin. However, IFN-gamma induced Ia antigens on only B cells, macrophages and T-dependent mast cells or persisting cells. On the basis of these results, we suggest that a major function of IFN-gamma is to potentiate immune responses by enhancing the expression of H-2 and Ia antigens on a variety of cell types.

Isotypes and specificities of immunoglobulins produced by germ-free mice fed chemically defined ultrafiltered "antigen-free" diet.

Abstract: The regulation of the “spontaneously” occurring (“background”) Ig synthesis of mice has been studied by determining the numbers of IgM-, IgG- and IgA-secreting cells and a part of the IgM antibody-specificity repertoire in spleen, bone marrow (BM) and mesenteric lymph nodes (MLN) of conventional and “antigen-free” mice. These antigen-free mice were germ-free raised and fed an ultrafiltered solution of chemically defined low molecular weight nutrients, and thus devoid of exogenous antigenic stimulation. The secretion of IgM, IgG and IgA by spleen, BM and MLN cells was assessed in the protein A plaque assay, while specific IgM antibody-secreting cells were detected by plaque assays specific for differently haptenated sheep red blood cells. In general, antigen-free and conventional mice were found to have roughly equal numbers of IgM-secreting cells in spleen and BM. The number of IgG-secreting cells in the spleen of antigen-free mice was the same as in the spleen of conventional mice, but in the BM their number was 3–5-fold decreased. About one half of the antigen-free mice did not have MLN, and in the half which did, 5 times less IgM- and more than 100-fold less IgG-secreting cells were found as compared with conventional mice. The number of IgA-secreting cells in antigen-free mice was drastically decreased in all three organs tested. The antibody-specificity repertoire of the “background” IgM-secreting cells in the spleen and BM of the antigen-free and conventional mice was much alike. This indicates that in antigen-free mice the available antibody repertoires are established independently of exogenous and/or mitogenic stimulation.

The origin of follicular dendritic cells in the mouse and the mechanism of trapping of immune complexes on them

Abstract: F1----parental bone marrow chimeras between CBA and B10 mice were used to show that even after 1 year follicular dendritic cells (FDC), also termed dendritic reticular cells, are not derived from bone marrow stem cells. However clusters of cells wrapped in the dendritic processes of FDC, isolated by enzyme digestion of the spleen contain B cells originating from donor marrow. This implies that evidence for the presence of antigens such as Ia-like detected by others (Gerdes, J. and Stein, H., Clin. Exp. Immunol. 1982. 48: 348) by immunohistology in or on FDC clusters should be interpreted with caution. Deposition of immune complexes on mouse spleen FDC depends upon the presence of a radio- and cyclophosphamide-sensitive population of cells, although FDC themselves are resistant to such treatments. After whole body irradiation the capacity to localize fluorescein isothiocyanate-labeled aggregated human gamma globulin can be restored by normal or T-deprived spleen cells, but restoration requires an interval of more than 1 though less than 5 days. These findings are compatible with the evidence of Gray et al., Eur. J. Immunol. 1984. 14: 47, that in the rat immune complexes are transported to FDC by a sessile population of marginal zone lymphocytes, as first suggested by Brown et al., Immunology 1973. 24: 955.

HLA-DC antigens can serve as recognition elements for human cytotoxic T lymphocytes.

Abstract: The specificity of four cytotoxic T lymphocyte (CTL) clones which recognize class II major histocompatibility complex (MHC) antigens was analyzed. All clones recognized antigens associated with the serologically defined HLA-DRw6 specificity. The activity of two of these clones, JR-2-2 and JR-2-10, could be inhibited by a monoclonal antibody Q 5/13 specific for a monomorphic determinant present on HLA-DR. In contrast, the activity of the two other CTL clones, JR-2-19 and JR-2-26, was not blocked by Q 5/13, but by a new monoclonal reagent, SPV-L3. This latter monoclonal antibody precipitated a two-chain structure of 28 kDa and 33 kDa and reacts with a monomorphic determinant. The molecular weight of the polypeptides precipitated with SPV-L3 was slightly less than those precipitated with a HLA-DR-specific monoclonal reagent. In addition two-dimensional gel electrophoresis showed that the antigen precipitated by SPV-L3 differed in charge from those precipitated with the anti-HLA-DR antibody. These results indicate that SPV-L3 recognizes a class II MHC product different from HLA-DR. This observation was confirmed by partial amino acid sequence analysis of the two chains which revealed that the molecule precipitated by SPV-L3 is homologous to HLA-DC/DS molecules. Therefore this report provides the first evidence that human cytotoxic T cells can recognize HLA-DC/DS antigens.

Thymocytes appear to ignore class I major histocompatibility complex antigens expressed on thymus epithelial cells

Abstract: BALB/c nu/nu mice were grafted with embryonic 14-day-old C57BL/6 thymi which were transplanted either nontreated or after elimination of hemopoietic cells with 2-deoxyguanosine. In both types of grafts host cells developed normally into functional thymocytes. Thymocytes from 2-deoxyguanosine-treated but not from untreated grafts contained as many cytolytic T lymphocyte precursors specific for class I MHC antigens on thymus epithelium as normal BALB/c thymocytes. As cytolytic T lymphocyte precursors were neither suppressed nor activated in these grafts it is concluded that thymocytes ignore class I MHC antigens expressed on thymus epithelium.

Cloned helper T cells can kill B lymphoma cells in the presence of specific antigen: Ia restriction and cognate vs. noncognate interactions in cytolysis.

Abstract: Cloned, Lyt-1+,2-, antigen-specific, Ia-restricted T cell lines can inhibit the growth of Ia-bearing B lymphoma cells in the presence of specific antigen. This effect is due to cytolysis of the B lymphoma cells in an antigen-specific, Ia-restricted manner by the cloned T cell lines. These cloned T cell lines can also kill lipopolysaccharide-activated normal B cells, while they activate resting B cells to divide and secrete immunoglobulin and are thus helper T cells as well as cytolytic T cells. The mechanism of cytolysis was examined in detail. Killing was mediated by a nonspecific mechanism after specific stimulation of the T cells with antigen presented in the context of the appropriate Ia glycoprotein complex, possibly implying a role for a soluble mediator. This simple system involving two clonal populations allows a detailed analysis of T-B interactions. Our studies are consistent with the view that both cognate and noncognate interactions of Ia-restricted T cells with B cells are mediated by nonspecific factors. Thus, the difference between interactions that appear to be cognate and those that appear to be noncognate may be quantitative rather than qualitative. That two cloned populations of cells can show either pattern of interaction depending on T-B ratio provides strong support for this view. Finally, that cloned helper T cells can kill activated B cells in an antigen-specific fashion may provide a new mechanism of immune regulation that would be especially important in responses to self antigens in vivo.

Inhibition of IgE binding to mast cells and basophils by monoclonal antibodies to murine IgE.

Abstract: In an attempt to identify the site on IgE which binds with high affinity to the Fc epsilon receptor (Fc epsilon R) on mast cells, we established monoclonal anti-IgE antibodies (mAb) by fusion of myeloma cells with rat splenocytes immunized with purified murine IgE mAb. Six individual mAb were found to react with various IgE mAb of different specificities and not with immunoglobulins of other classes. Three different clusters of epitopes on the Fc epsilon portion could be detected by antibody competition studies. These antigenic determinants were expressed on the Fc epsilon portion and required the two heavy chains in their native conformation. Two groups of mAb and their Fab' fragments completely inhibited the binding of 125I-labeled IgE to rat basophilic leukemia cells (RBL), and one mAb inhibited the specific IgE binding only partially (55-65%). Likewise, the Fab' fragments of the purified mAb inhibited the antigen-mediated, IgE-dependent, serotonin release of RBL cells. These in vitro findings were confirmed by in vivo experiments, which demonstrated that the anti-IgE mAb could specifically block passive cutaneous anaphylaxis reaction when injected i.d., before challenging with the antigen. The differences in blocking reactivity of the various anti-IgE mAb are discussed in view of heterogeneity in the IgE-Fc epsilon R interaction.

Characterization of human granular lymphocyte subpopulations expressing HNK-1 (Leu-7) and Leu-11 antigens in the blood and lymphoid tissues from fetuses, neonates and adults.

Abstract: Three subpopulations of human granular lymphocytes from blood and lymphoid tissues were characterized using combinations of the monoclonal antibodies (mAb) HNK-1 (Leu-7), Leu-11 and VEP13. Each subpopulation was confirmed to possess natural killer (NK) cell functional capability, but a different level of cytotoxic efficiency (HNK-1+leu-11- less than HNK-1+Leu-11+ less than HNK-1-Leu-11+). In adult from 23 healthy donors, the subpopulations with HNK-1+Leu-11-, HNK-1+Leu-11+ and HNK-1-Leu-11+ phenotypes comprised 4.7 +/- 3.0, 8.0 +/- 6.4 and 3.9 +/- 3.5% of mononuclear cells, respectively. Despite their distinct surface marker phenotypes and NK functional ability, all 3 subpopulations exhibited granular lymphocyte morphology. One of these subpopulations, HNK-1+Leu-11-, also expressed the pan-T cell antigen Leu-4. Different patterns were observed in fetal bone marrow and cord blood, where the vast majority of HNK-1+ cells lacked the Leu-11 antigen (HNK-1+Leu-11+ cells). The HNK-1 antigen was not expressed on granulocytes and their precursors, whereas both Leu-11 and VEP13 antigens were expressed on these myeloid cells from fetal bone marrow and cord blood as well as adult bone marrow and spleen. Cell lines of granular lymphocytes cultured in the presence of interleukin 2 all possessed the HNK-1+Leu-4+ phenotype and NK functional capability but lacked the Leu-11 and VEP13 antigens on their surface after 15 days of culture. Although granular lymphocytes expressing the Fc receptors reacting with the mAb Leu-11 and VEP13, are the most functionally active NK cells, the HNK-1+ subpopulation lacking the Leu-11 and VEP13 antigens appears to be an important population (possibly an immature form of granular lymphocytes) for delineating the cell lineage(s) and differentiation of human granular lymphocytes. Although none of the currently available mAb react both inclusively and exclusively with human granular lymphocytes, the combination usage of these antibodies permits a more precise and comprehensive analysis of these subsets.

Monoclonal anti-cardiolipin antibodies bind to DNA.

Abstract: BALB/c mice immunized with the phospholipid, cardiolipin, produced anti-cardiolipin and anti-DNA antibodies. Seven hybridomas derived from spleen cells of the cardiolipin-immunized mice produced cardiolipin-binding monoclonal antibodies that also bound to the polynucleotides DNA, poly(dT), and poly(I). The seven cardiolipin-induced monoclonal antibodies shared idiotypic determinants with a high frequency idiotypic marker of spontaneously expressed anti-DNA autoantibodies of lupus-prone MRL-lpr/lpr mice. The monoclonal antibodies presumably bound to phosphodiester phosphate groups that occur in both polynucleotides and phospholipids. The results imply that production of anti-DNA autoantibodies does not require immunization by DNA.

Pure interferon gamma enhances class II HLA antigens on human monocyte cell lines.

Abstract: Human recombinant interferon gamma (IFN-gamma) with a chemical purity of over 90% was shown to enhance membrane expression of HLA-DR antigens on cells from 3 human myelomonocytic lines (HL-60, U-937 and THP-1). Immunofluorescence techniques using a series of anti-HLA-DR monoclonal antibodies showed that the low, although variable, levels of DR-positive cells were clearly enhanced as soon as 24 h after incubation with IFN-gamma. Only IFN-gamma was able to exert this effect, since incubation with high concentrations of IFN-alpha or -beta did not induce any significant modification of the percentage of HLA-DR-positive cells. In contrast, doses of IFN-gamma as low as 2 units were effective, indicating a highly preferential, apparently selective effect of IFN-gamma for enhancement of HLA-DR expression. Private class II antigen expression was also enhanced by IFN-gamma on the U-937 cell line. Through its enhancing effect on class II public and private HLA antigens on the membrane of human monocytes, the IFN-gamma lymphokine may have a critical role in the modulation of antigen presentation by monocytes and on the regulation of HLA-DR-restricted cell cooperation.

Identification of an anti‐monocyte monoclonal antibody that is specific for membrane complement receptor type one (CR1)

Abstract: A monoclonal antibody (E11) was produced by immunization of mice with intact human cells of monocyte lineage. Despite the finding that E11 did not inhibit rosettes with C3b-coated sheep erythrocytes (EC3b), several lines of evidence indicated that E11 was specific for complement receptor type one (CR1). All monocytes, neutrophils, lymphocytes and erythrocytes that reacted with E11 formed EC3b rosettes. The E11 antigen on these cells was shown to be a molecule of 222 +/- 10 kDa. Treatment of lymphocytes, monocytes, and neutrophils with E11 followed by fluorescein-coupled F(ab')2 anti-mouse-IgG at 37 degrees C in buffer lacking sodium azide, led to capping or apparent endocytosis of the E11 antigen and a diminution in CR1 activity of 88%, 59% and 25%, respectively. This same treatment had no detectable effect on monocyte or neutrophil CR3 activity (EC3bi rosettes). Furthermore, with E11-capped lymphocytes, the residual EC3b rosetting was capped directly over the E11-fluorescence cap, whereas EC3d,g rosetting (CR2 specific) was undiminished and distributed evenly around the circumference of cells containing E11-fluorescence caps. Finally, the binding of E11 to cells was inhibited by the prior treatment of these cells with a well characterized rabbit polyclonal anti-CR1. These data indicated that E11 was specific for a site in CR1 that was distal from the C3b-binding site, so that E11 was unable to block CR1 activity. E11 proved to be useful for identifying CR1 on various cells in tissue sections, and for quantitating CR1 on erythrocytes and neutrophils. Erythrocytes and neutrophils from normal individuals were found to bind an average of 610 and 4.6 X 10(4) 125I-labeled E11 molecules per cell. When E11 was visualized in tissues by immunoperoxidase staining, the cells that apparently contained the greatest amounts of CR1 were dendritic reticulum cells and kidney podocytes. The E11 reactive dentritic reticulum cells were characteristic of both follicular and diffuse follicular center cell tumors. Lymphocytes from patients with chronic lymphocytic leukemia (CLL) characteristically expressed little E11, confirming earlier studies that CLL cells lacked CR1 activity detected by EAC1-3b rosette formation. Because normal B cells have been shown to express CR1 at a very early stage of maturation, the absence of CR1 on CLL cells is discordant with the immature nature of CLL cells defined by immunoglobulin expression.

The cytolytic T lymphocyte response to the murine cytomegalovirus. II. Detection of virus replication stage‐specific antigens by separate populations of in vivo active cytolytic T lymphocyte precursors

Abstract: During the acute cytolytic T lymphocyte (CTL) response of mice to infection with the murine cytomegalovirus two independent populations of activated interleukin-receptive CTL precursors can be demonstrated. One population is specific for cell membrane-incorporated viral structural antigens, whereas the second population detects an antigen, whose appearance is correlated with the synthesis of viral immediate early proteins. Since this new type of antigen is only defined by lymphocyte recognition, it is referred to as the lymphocyte-detected immediate early antigen (LYDIEA). Expression of immediate early antigen precedes the production of viral progeny and, therefore, it is possible that LYDIEA-specific CTL could serve as indicator cells for the very first activities of the viral genome, even during nonproductive infection.

Both Fc receptors and lymphocyte-function-associated antigen 1 on human T gamma lymphocytes are required for antibody-dependent cellular cytotoxicity (killer cell activity).

Abstract: A monoclonal antibody, designated CLB-LFA-1/1, directed to the human lymphocyte-function-associated antigen 1 (LFA-1) was raised by immunization of mice with the peripheral blood lymphocytes of a T gamma lymphocytosis patient. The monoclonal antibody was selected by inhibition of the natural killer cell and the antibody-dependent killer cell activity of the patient's T gamma lymphocytes. In addition, the monoclonal antibody was shown to inhibit the cytotoxic activity of T cell clones specific for either class I or class II HLA molecules. The antigen recognized by CLB-LFA-1/1 consisted of three polypeptide chains with molecular weights of 180 000 (alpha), 155 000 and 94 000 (beta). The antibody reacted with T cells, B cells, monocytes and granulocytes, and stained normal T gamma cells and T gamma cells of patients with T gamma lymphocytosis two- to threefold stronger than normal T cells. It was shown that LFA-1 and the Fc receptor on T gamma cells did not comodulate and it is therefore concluded that Fc receptors and LFA-1 are independent membrane structures, both required for the killer cell activity of T gamma cells.

The effect of antigen on the development of Peyer's patches in sheep

Abstract: The proposition that the development of Peyer's patches (PP) is influenced by anti-genic stimulation has been examined in sheep. Terminal lengths of ileum containing about half of the ileocecal PP were isolated from the intestinal tracts of fetal lambs during the last month before birth. Antigen was injected into some of these segments and the subsequent development of the PP studied before and after birth. The injection of either killed B. abortus, ferritin or maternal colostrum into the lumens of the isolated ileal segments did not cause premature growth of the PP follicles, nor did it effect the content of lymphoblasts in them. In contrast, the injection of these antigens into the isolated segments caused the development of germinal centers and plasma cells in the regional mesenteric lymph nodes. Plasma cells also appeared in the lamina propria along the intestinal tract in response to these antigens. These results provided experimental evidence that lymphopoiesis in the follicles of the PP of fetal lambs is not dependent on antigen. The PP in ileal segments that were not injected with antigen and had no contact with antigen subsequently grew at the normal rate before and for the first 2 weeks after birth; after this the growth of the follicles became significantly slower than normal. The follicles in these isolated ileal segments had almost completely disappeared by 3–4 months of age, whereas the follicles in the normal functional ileum did not undergo involution until around 15 months of age. The premature involution in the PP in the isolated segments was prevented by reconnecting the segment to the functional intestinal tract before 3 months of age.

Induction of resting B cells to DNA synthesis by soluble monoclonal anti-immunoglobulin.

Abstract: A novel monoclonal anti-immunoglobulin is described which in soluble form is comparable to lipopolysaccharide in its ability to induce DNA synthesis in populations of resting B cells. It inhibits lipopolysaccharide-induced B cell maturation to immuno-globulin secretion while not suppressing mitogen-induced proliferative responses. B cells from C3H/HeJ mice are stimulated as well as those from C3H/Tif mice demonstrating that the induction capacity of this monoclonal antibody does not involve functional lipopolysaccharide receptor molecules.

Functional significance of the regulation of macrophage Ia expression.

Abstract: The relationship of Ia expression and antigen-presenting function by macrophages has been evaluated. When macrophages are maintained in standard culture media, both Ia antigens and accessory cell function are lost. The reacquisition of these properties follows exposure to an Ia-inducing lymphokine, for which cDNA-derived interferon-gamma may substitute. The induction of function is related quantitatively to the level of Ia expression. Moreover, both properties reflect newly expressed Ia determinants, since treatment with anti-I-A plus complement at the beginning of culture diminishes neither the subsequent level of Ia expression nor function. Treatment with anti-Mac-1 plus complement, however, reduces function commensurate with the effectiveness of macrophage depletion. Finally, we find that fixation of macrophages after exposure to antigen does not inhibit antigen presentation, indicating that metabolic activity, while required for antigen processing, is not necessary for presentation.

Monoclonal antibodies specific for murine IgM. I: Characterization of antigenic determinants on the four constant domains of the μ heavy chain

Abstract: Seventeen monoclonal rat antibodies with specificities for mouse mu heavy chain recognize seven distinguishable determinants that are located on the four constant region domains. All determinants are present on secreted, intracellular and membrane bound IgM, and all but two are expressed on isolated mu heavy chain. One antibody, specific for a site in the first constant region domain, recognizes a determinant that is present on IgM of AKR, C3H/HeJ, C57BL/6J, SJL, DBA/2, BALB/c, NZB and CBA/J mouse strains, but not on IgM of A.TH, A/J and A.CA strains of mice.

Thymic hormone-containing cells. VI: Immunohistologic evidence for the simultaneous presence of thymulin, thymopoietin and thymosin α1 in normal and pathological human thymuses

Abstract: The localization of the three best-defined thymic hormones, namely, thymulin, thymopoietin and thymosin alpha 1 was studied by immunofluorescence using antibodies directed against these three molecules. With both human thymus frozen sections and cultured cells, thymic hormones were found exclusively in the epithelial component (recognized by its keratin content), in normal as well as pathological thymuses. The double-labeling experiments using the different anti-thymic hormone antibodies showed that the same epithelial cells contained the three hormones. These results suggest that the production of different hormones in the thymus is accomplished by the same epithelial cells.

The percentages of six immunoglobulin isotypes in human antibodies to tetanus toxoid: Standardization of isotype‐specific second antibodies in solid‐phase assay

Abstract: The solid-phase radioimmunoassay for human antibodies was improved so that it gave the total concentration as well as the concentrations of IgM, IgA, IgG1, IgG2, IgG3 and IgG4 antibodies, all seven in the same "L units"/ml. This was accomplished by standardization of monoclonal antibodies with monoisotypic antibodies rather than myelomas; myelomas were found unsatisfactory for this purpose. For the first time it was possible to determine the isotype composition of an antibody population in percent terms. The weight equivalent of the L-unit in one hyperimmune tetanus antitoxin preparation was approximately 7.4 ng. The new solid-phase radioimmunoassay was applied to tetanus toxoid antibodies of the booster response. Total concentrations varied from 1700-26 000 L units/ml (13-200 micrograms/ml). Concentrations of IgG1 antibodies were 1600-25 000 units/ml (average 91% of the total) and concentrations of IgG4 antibodies 12-6900 units/ml (average 6.9% of the total). Antibodies of the other four isotypes were not detected in all sera and together they never exceeded 3% of the total.

Synergistic protection by specific antibodies and interferon against infection by Trypanosoma cruzi in vitro

Abstract: Trypanosoma cruzi infects many mammalian cell types in vitro, including fibroblasts and macrophages. We have analyzed and compared the infection of mouse 3T3 fibroblasts and J774 macrophage-like tumor cells by cloned Y-A1.2 T. cruzi trypomastigotes. The effects of T. cruzi-specific antibodies and of interferon (IFN) on infection were considered. Specific antibodies protected 3T3 fibroblasts from infection by T. cruzi, but increased the relative infectivity for J774 cells due to binding of the opsonized parasite to J774 Fc receptors. It was also apparent that IFN-alpha, beta (produced by fibroblasts) and IFN-gamma (produced by T lymphocytes) activated trypanocidal activities in both 3T3 and J774 cells, although IFN-gamma was 100 to 2000 times more effective than IFN-alpha, beta on the basis of IFN antiviral units added per culture. When anti-T. cruzi antibodies and IFN-alpha, beta or IFN-gamma were used jointly, a synergistic protective effect was observed with both 3T3 fibroblasts and J774 cells. These results suggest that B and T lymphocytes might collaborate in vivo in the protection against T. cruzi infections by the production of anti-trypomastigote antibodies and IFN-gamma, respectively.

Chicken thymocyte-specific antigen identified by monoclonal antibodies: ontogeny, tissue distribution and biochemical characterization

Abstract: Two monoclonal antibodies, CT-1 (IgG1, kappa) and CT-1a (IgG3, kappa) were prepared against chicken thymocytes. The antigen identified by these antibodies was found to be a glycoprotein with a major polypeptide component having an apparent molecular weight of 63 000 and a minor polypeptide component of approximately 103 000. Immunoprecipitation and blocking experiments revealed that the two antibodies react with different antigenic determinants of the molecule. Ontogenic studies employing immunofluorescence failed to reveal the antigenic determinants on cells from the embryo or embryonic yolk sac on days 3 and 6 of incubation. The number of embryonic thymocytes bearing the molecules detected by these antibodies increased from 6% on day 12 to 54% on day 13; the frequency of thymocytes expressing this glycoprotein reached adult levels (greater than 90%) by the 15th day of embryonic age. In contrast, the CT-1 and CT-1a antibodies reacted with only 2-5% of blood and splenic cells and less than 0.05% of cells from bursa or bone marrow of young adult chickens. In the quail CT-1 reacted with cortical, but not medullary, thymocytes, while the CT-1a antibody was unreactive with quail thymocytes. The surface glycoproteins detected by these discriminating monoclonal antibodies may provide an important discriminating marker for thymic lymphocytes in the chicken and the quail.

Isolation of follicular dendritic cells from human tonsils and adenoids. II. Immunocytochemical characterization.

Abstract: Follicular dendritic cells (FDC) are specialized cells found only within lymphoid follicles. They bind immune complexes and play a role in the presentation of antigen to follicular B cells and in the generation of B cell memory. In the present report the isolation of FDC from human tonsils and adenoids is described. These isolated cells have an unusual spherical arrangement and enclose lymphocytes within extensions of their membranes. Their ultrastructural features are similar to those observed in situ. The reactivity of isolated FDC with a number of monoclonal antibodies was analyzed by immunofluorescence and by immunostaining (at the electron microscopic level) with colloidal gold. In keeping with the results of previous investigations on tissue sections IgM, IgG and IgA (but not IgD) can be detected on the surface of isolated FDC, as can C3b receptors and the FDC-associated antigen detected by monoclonal antibody R4/23. The immunoglobulins associated with FDC are mostly embedded in an electron-dense material. The majority of the lymphoid cells enclosed within the membrane extensions of FDC are of B cell type. These results suggest that isolated FDC may be suitable for further in vitro investigation of their role in the humoral immune response.

Autoregulation of autoantibody synthesis in mercuric chloride nephritis in the Brown Norway rat. I. A role for T suppressor cells.

Abstract: Mercuric chloride injections in the Brown Norway rat induce the transient formation of anti-glomerular basement membrane (GBM) autoantibodies. Transfer of spleen cells from convalescent animals, after circulating anti-GBM autoantibodies are no longer detectable, inhibits reinduction of the disease by HgCl2 in naive recipients. This inhibition is significantly less when the T suppressor cell population is depleted by the monoclonal antibody, MRC OX8 , before transfer. Our studies suggest a role for T suppressor cells in autoregulation in this animal model of autoimmune nephritis and may form a basis for the design of specific therapy for anti-GBM disease in man.

Different mechanisms of release of vasoactive amines by mast cells occur in T cell-dependent compared to IgE-dependent cutaneous hypersensitivity responses.

Abstract: It has recently been shown that the classical 24-48-h component of delayed-type hypersensitivity (DTH) in mice is preceded by an early 2-h component. This early component of DTH resembles IgE-dependent hypersensitivity reactions, but is mediated by an antigen-specific T cell factor that activates cutaneous mast cells (MC). In the current study of cutaneous hypersensitivity reactions, evidence is presented that serotonin and histamine are released from the granules of MC in IgE-dependent responses, but that T cell activation of MC induces preferential release of serotonin. In T cell-dependent reactions, it was found that serotonin was released under experimental conditions in which MC granules were depleted of serotonin, but catabolism of serotonin in the cytosol was prevented. This suggests that T cells induce differential release of serotonin from non-granule storage sites in MC, such as transport vesicles in the cytoplasm. Morphological observations of MC in IgE-dependent compared to T cell-dependent cutaneous reactions were consistent with this hypothesis. Exocytosis of granules characterized IgE activation and was not found in MC that were activated by T cells, which instead showed mild degranulation accompanied by numerous cytoplasmic vesicles.