A monoclonal antibody to the HLA-A3 alloantigen

Monoclonal antibody production recognizing the HLA-A3 antigen is described. The XI-23 antibody reacted with all of the 89 cell suspensions carrying the HLA-A3 antigen (100% cytotoxicity) among a total of 191 suspensions tested. No extra-reactivity or cross-reactivity was observed, particularly with that of HLA-A11. This antibody can thus be considered as a good HLA-typing reagent.     

A human monoclonal antibody against HLA-A25

We are reporting the production and characterization of a human monoclonal antibody recognizing antigen HLA-25. The antibody was developed by a line transformed in vitro by the Epstein-Barr virus. The immune B lymphocytes for transformation were generated by planned immunization of a volunteer with repeated doses of allogenic peripheral blood lymphocytes of one donor over the course of 7 years. The antibody showed correlation with A25 antigen on a panel of 244 individuals tested by microcytotoxicity. The antibody showed neither cytotoxic reactivity nor CYNAP phenomenon with antigens of HLA-10 CREG.     

A monoclonal antibody against HLA-A11 and A24

A monoclonal IgG antibody was produced from a mouse immunized with an A11, A24; B27, B44 Epstein-Barr virus transformed B lymphoblastoid cell line. The antibody, A11.1M, by standard lymphocytotoxicity assay, reacts with all cells expressing HLA-A11 and -A24. Absorption studies with both A11+, A24- and A11-, A24+ platelets removed antibody reactivity against A11 and A24 lymphocytes. The shared antigenic determinant between A11 and A24, as defined by this antibody, A11.1M, represents a new "supertypic" determinant.     

A monoclonal antibody detecting an HLA-DQwl-related determinant

A complement fixing monoclonal antibody (moab) was prepared which reacts with a polymorphic determinant on HLA class II molecules. The moab IIB3 recognises all DQwl (DC1, MB1, LB-E12) positive cells as well as some DR4, DR7, DRw8 and DRw9 positive cells. The moab reacts mainly with B-cells and not or with only a minority of the monocytes. Segregation of the determinant with HLA-DR could be shown. The determinant is strongly expressed on DR2, DR4 and DRw6 positive cell lines but only weakly on DR1 lines. In contrast to a monoclonal antibody against a monomorphic determinant on class II molecules IIB3 did not give a distinct inhibition of the MLC nor did it inhibit the generation of CTLs in MLC as has been described for the DQwl like moab BT 3/4 (Corte et al. 1982). Immunoprecipitation indicates that IIB3 reacts with DQ-like molecules.     

Two monoclonal antibodies detecting allotypic determinants of HLA-A

Three mouse hybridomas producing cytotoxic antibodies against HLA were established. By standard microcytotoxicity test against panels of normal controls, the antigen defined by MA-9 antibody (IgM) showed a good correlation with HLA-A9 alloantigen detected by conventional typing alloantisera (r = 1.0). Family studies also showed that MA-9 determinant segregated with HLA-A9. MA-10 antibody (IgM) reacted with all HLA-A10 positive lymphocyte donors and cross-reacted with two thirds of HLA-AW33 positive donors. M1-1 antibody (IgG2a) reacted with all the panel cells tested and immunoprecipitated a molecule of 43,000 daltons from Nonidet P-40 lysates of ³H-glucosamine-labelled cells. The results showed that MA-9 and MA-10 antibodies can be used as routine tissue typing reagents.     

A mouse monoclonal antibody to the TSHR

INTRODUCTION: Mouse monoclonal antibodies (mAbs) with the ability to inhibit thyrotropin (TSH) binding to the TSH receptor (TSHR) are useful tools to study TSH-TSHR interaction. The 3C3 mAb we produced was found to inhibit binding of TSH to human (h)TSHR but not to porcine (p)TSHR. MATERIAL/METHODS: Purified 3C3 immunoglobulin G (IgG) and its antibody-binding fragment were prepared using standard methods and their ability to inhibit TSH binding to hTSHR or pTSHR was analyzed using a coated tube assay. The TSHR epitope reactive with 3C3 IgG was determined using Western blotting, ELISA based on peptides corresponding to the TSHR sequence, and the SPOT synthesis technique. RNA was isolated from 3C3 hybridoma cells and the mAb variable (V) region genes were sequenced and analyzed. RESULTS: 3C3 mAb had a 1 x 108 l/mol binding affinity to the hTSHR as assessed by Scatchard analysis. 3C3 reacted with the hTSHR region between amino acids (aa) 212-230, and two aa differences were found between the corresponding regions in the hTSHR and pTSHR. The light chain (LC) genes of 3C3 were derived from the Vk21 germ-line (97.6% homology) and Jk2 genes. The heavy chain (HC) genes were from the V130 germ-line (94.6% homology) combined with a D gene (not identified) and JH3 gene. The replacement/ silent mutation ratios of 6.0 and 6.5 for the LC and the HC V regions, respectively, indicated that 3C3 underwent antigen-driven maturation. CONCLUSIONS: Mouse mAbs of this type should be useful in studying the interactions between the TSHR, TSH, and mAbs in more detail.     

A mouse monoclonal antibody detecting a DR-related MT2-like specificity: Serology and biochemistry

A mouse monoclonal antibody (7.3.19.1) was produced which reacts with class II molecules on B cells and monocytes of DR3, DR5, and/or DRw6 positive donors only. Using this moab and two others, three different groups of class II molecules could be identified. Furthermore, a differential precipitation pattern was found which correlates with a DR-related variable expression of the MT2-like polymorphic determinant on the cell surface. Addition of 7.3.19.1 to MLCs did not result in significant inhibition in contrast to the two other moabs tested. Normal CTL activity was found in such a stimulated responder population.     

H9/25 monoclonal antibody recognizes a new allospecificity of mouse lymphocyte subpopulations: Strain and tissue distribution

C3H/He-mg mice were immunized with C57 BL/10 (B10) spleen cells and the immune spleen cells were fused with BALB/c myeloma cells (NS1). One of the monoclonal antibodies (H9/25 antibody) produced by the hybrid cells was studied. It reacts with subpopulations of B 10 lymphocytes as well as some lymphoid tumor lines including some of the Abelson virus-induced leukemias. The antigen recognized by H 9/25 antibody is expressed on lymphocytes from all the B 10 congeneic mice tested as well as some other strains of mice. No linkage between genes coding for the antigen and H-2 loci was found as judged by its presence on cells of the B 10 strains regardless of H-2 type and the distribution of the antigen on Bailey recombinant inbred mice. The antigen is expressed on subpopulations of lymph node cells, spleen cells, thymocytes and bone marrow cells. The strain distribution of the H9/25 antigen seems to be identical to that of Ly-6, Ly-8 and Ala-1 antigens. However, the tissue distribution of the antigen recognized by H9/25 antibody, while similar to these alloantigens, is unique and the antigen may be distinct from the other alloantigens.     

A monoclonal antibody detecting dipeptidylpeptidase IV in human tissue

Summary Dipeptidylpeptidase IV (DPP IV) occurs among others in exocrine epithelia, hepatocytes, renal tubuli, endothelia, and myofibroblasts of man and laboratory animals. Also Tµ lymphocytes and their varying differentiated neoplastic counterparts reveal this enzyme activity. The present paper describes a new monoclonal antibody recognizing DPP IV.Additional efforts have been taken to detect the subcellular localization of DPP IV and its isoelectric focusing pattern in different tissue types. The monoclonal antibody anti-DPP IV (clone II-19) shows a reaction pattern indistinguishable from the corresponding enzymehistochemical reaction. These findings were further substantiated by immunoblotting analysis. In line with the results of direct enzyme measurements in different subcellular fractions a considerable portion of the enzyme is localized in the membrane fraction.Dedicated to Professor Dr.Dr. h.c. Karl Lennert, Kiel, on the occasion of his 65th birthdayThis study was supported by the Deutsche Forschungsgemeinschaft, SFB 111, program CL1     

识别人白细胞免疫标志的单克隆抗体

A monoclonal antibody (48)against human Pan-Leucocytes was prepared. This antibody reacted with all haemopoietic cells tested, but not with red cells and platelets by indirect immunofluorescent staining. It was also disclosed that this antibody is only bound to lymphoid tissues or leucocytes scattered in other tissues with immunoperoxidase staining. The reactivities of McAb 48 with large a mount of various target cells were identical with anti-HLE, McAb of CD45 group. Therefore McAb 48 also recognizes T200 antigen and belongs to CD45 group. The value of McAb 48 in differential diagnosis of malignant lymphoma is discussed.     

A cytotoxic anti HLA-AB monoclonal antibody which in dilution becomes specific to HLA-A3 crossreacting group

XI 20.4 monoclonal antibody belongs to the IgM class. It precipitates two polypeptide chains characteristic of HLA Class I antigens. At the highest dilutions it is cytotoxic against lymphocytes carrying antigens of the HLA-A3 crossreacting group. Lysostrip experiments show that, at the lowest dilutions, the antibody reacts either with HLA-A, or B antigens.     

Monoclonal antibody to HLA-A3

The monoclonal antibody GAP A3 detects the HLA allospecificity A3. Reactivity of the monoclonal was in exact concordance with the presence of the A3 antigen as defined by conventional alloantisera on a panel of 59 cells from individuals with well-characterized HLA antigens. Reactivity with GAP A3 segregated with HLA-A3 in a family where three of eight siblings inherited the paternal A3 antigen. GAP A3 precipitated appropriate 44,000- and 12,000-dalton bands on SDS-polyacrylamide gels under reducing conditions from an HLA-A3-positive, but not an HLA-A3-negative B lymphoblastoid cell line. Thus, by serological, familial, and biochemical criteria, GAP A3 defines the allospecificity HLA-A3.     

Generation and characterization of three new monoclonal antibodies detecting the allospecificities HLA-A2, w69, HLA-A3 and HLA-B13

Immunization of balb/c mice with peripheral blood mononuclear cells of a leukemic patient possessing the antigens HLA-A2,3; B7,35 resulted in the polymorphic monoclonal antibody (moab) UL-101/68 (IgM) defining HLA-A3. Immunization of a second group of balb/c mice with the lymphoblastoid cell line BER homozygous for HLA-A2; B13 revealed two polymorphic moabs UL-39/10 (IgG3) specific for HLA-B13 and UL-39/24 (IgG2b) defining HLA-A2,w69. Immunoprecipitation and polyacrylamide gel electrophoresis with the two moabs of the IgG isotype confirmed the class I structure of the recognized antigens.     

A monoclonal antibody detecting a possible public specificity of the HLA-C locus

An IgM monoclonal antibody (CC-Cl 11) was produced by fusing myeloma cell line SP2/08 with lymphocytes of a Balb/c mouse previously immunized with peripheral blood lymphocytes of an A2, Bw44, B27, Cw1, Cw7, DR5 donor. Reactivity of CC-Cl 11 on a lymphocyte panel of 172 unrelated donors and lysostripping and absorption experiments have shown that CC-Cl 11 recognizes an antigenic determinant common to HLA-Cw1 and Cw3 positive lymphocytes.     

Description of a mouse monoclonal anti-HLA-B27 antibody HLA-ABC-m3

The production and characterization of a new anti-HLA-B27 monoclonal antibody HLA-ABC-m3 is described. This cytotoxic IgG2a antibody binds protein A and is able to precipitate cell surface molecules of 43,000 and 12,000 daltons corresponding to the HLA heavy chain and β2-microglobulin. Population testing revealed that the HLA-ABC-m3 antibody reacted with the peripheral blood lymphocytes of 47/47 individuals conventionally typed as HLA-B27⁺ and with 5/105 HLA-B27⁻ individuals. These five extra reactions were with individuals expressing the cross-reactive HLA-B7 alloantigen, although the affinity of the monoclonal antibody for B27 heterozygous individuals (approx 10⁹ M⁻¹) was tenfold greater than with B7 individuals (approx 10⁸ M⁻¹). In addition, HLA-ABC-m3 reactivity segregated with HLA-B27 in two families. This monoclonal antibody should be of value in the investigation of the role of HLA-B27 in disease.     

Immune and nonimmune effector functions of IgG3 mouse monoclonal antibody R24 detecting the disialoganglioside GD3 on the surface of melanoma cells

R24, an IgG3 mouse monoclonal antibody reactive with the disialoganglioside GD3, was found to be a potent mediator of human complement cytotoxicity and human effector cell cytotoxicity. Cytotoxicity correlated with the degree of antibody binding (GD3 cell surface expression) for each of the melanoma cell lines and melanocyte cell cultures tested. Melanoma cell lines binding low amounts of R24 (low GD3 cell surface expressors) were not lysed in R24-directed immune reactions, suggesting that a threshold number of R24 molecules bound per cell is necessary to initiate these cytotoxic mechanisms. Since both complement- and cell-mediated reactions lysed the same subpopulations of cells in each cell line, both mechanisms appeared to depend on similar threshold quantities of bound R24 molecules. However, due to the heterogeneity of R24 binding in each cell line, the numerical value for this threshold could not be determined. Only in cell lines binding greater than 10(7) R24 molecules per cell were greater than 90% of the cells lysed. Normal melanocytes in culture were not lysed by R24-directed immune mechanisms, due to their low GD3 expression, indicating that monoclonal antibodies such as R24 may show tumor specificity with regard to effector functions even though normal cells express the relevant antigen. In contrast to the potent in vitro activity of R24, treatment of nu/nu mice bearing human melanoma grafts resulted in tumor inhibition only when started within 3 days of tumor cell inoculation. No effect was seen on established tumors. Thus, this in vivo mouse model failed to predict the clinical and pathological findings observed in treatment trials of R24 in human melanoma patients--urticaria involving skin metastases, cellular infiltration of tumor tissue, and tumor regression. In addition to activating immunologic effector functions, R24 had direct effects on melanoma cells, blocking their ability to attach to surfaces and causing tumor cell aggregation. These effects were again related to the number of R24 molecules bound to the cell surface; no aggregation was seen with cell lines binding less than 4 X 10(5) molecules per cell. Both immune and nonimmune effector functions may be involved in the tumor inhibitory activity of R24 in humans.     

A mouse monoclonal antibody with HLA-DR4 associated specificity

A mouse monoclonal antibody is described which reacts with HLA-DR4 positive cells. Gel analysis of the immunoprecipitated antigen is consistent with it being a DR molecule.     

A monoclonal anti-HLA antibody recognizes a mouse tumor-associated antigen

The monoclonal antibody W6/32.1 recognizes a public determinant on the HLA-A, B and C antigens of all tested human haplotypes. Though the antibody does not bind to normal mouse cells of any H-2 haplotype, it does show an unexpected specificity for the T cell leukemia line MBL-2 from a C57BL/6 mouse. It is shown that the murine antigen recognized by W6/32.1 is on an H-2-like molecule which also carries the determinant recognized by the monoclonal antibody B22-249 R1, specific for the H-2Db antigen. Unlike B22-249 R1, however, W6/32.1 does not bind to normal H-2b lymphocytes, nor to a variety of tumor cell lines of the H-2b haplotype. This cross-reaction is specific to W6/32.1, and is not shared by other monoclonal antibodies of similar anti-HLA specificities. Moreover, the affinity of W6/32.1 for its human antigen is substantially higher than for its mouse antigen. We conclude that W6/32.1 fortuitously recognizes a novel determinant on the H-2Db antigen of MBL-2, rather than an extensive region of structural homology shared between HLA and H-2. Thus for cells of the H-2b allotype this determinant is detected only on MBL-2, and by definition is thus an example of a tumor-associated antigen.