摘要:

We have recently described a class-switched (IgM to IgG1) human-mouse chimeric antibody. In the present study, a human-mouse chimeric antibody specific for human adenocarcinoma-assocated atigen YH206 antigen was costrcted by fusing murine varible region genes (Vκ and VH) to human cnstant region genes (γ1,κ) The murine variable domain genes were isolated from a funconal murine hybridoma cell line, YH206, which secreted IgM monoclonal antibody specific for YH206 antgen. The fusion gens of heavy and light chins were introduced into the immunoglobulin non-prodcing mouse myeloma cell line X63-Ag8.653 by electroporation. We obtained transformants which secreted class-switched human-mouse chimeric antibodies specific for YH206 antigen. A dot immunobinding assay demonstrated that the class-switched chimeric antibody retained the ability to bind to the YH206 antig

ISSN:0272-457X    

DOI:10.1089/hyb.1991.10.1

年代:1991

数据来源: MAL

摘要:

We report the construction of a mouse-human (M-H) heterohybridoma by fusion of the murine myeloma cell line NS-1 and human spleen cells from a 17 week old fetus. The nonsecreting, cloned hybridoma cell line II was resistant to 8-azaguanine (8-AG) and sensitive to hypoxanthine, aminopterin and thymidine (HAT) medium. It grew rapidly in 8-AG containing medium (doubling time 20 hrs.), but did not grow in HAT medium or in non-serum medium. It had a high fusion frequency with human lymphocytes from regional lymph nodes. Five human chromosomes were retained stably for over 6 months by this cell line II. Nine (mouse-human)-human ((M-H)-H) triple hybridomas secreting human IgG 1 or IgM were established by the fusion of this parental cell line II and human lymphocytes from regional lymph nodes. Immunoglobulin secretion was stable and has been maintained for over 8-10 months without recloning in these hybridomas. Secretion of immunoglobulin varies from 2.1-3.0 μg/106cells/day, and these hybridomas contain from 3 to 16 human chromosomes, including No. 14. So, this M-H heterohybridoma II is an excellent useful parental cell line for the production of hybridomas secreting human immunoglobulin

ISSN:0272-457X    

DOI:10.1089/hyb.1991.10.11

年代:1991

数据来源: MAL

摘要:

Ahuman monoclonal antibody designated AC6C3 was developed by fusing regional lymph node lymphocytes from a patient with epithelial ovarian carcinoma with cells of the hybrid myeloma SPAZ 4. This monoclonal antibody recognized a determinant expressed on the cell surface of ovarian tumor cell lines. The AC6C3 hybridoma has been maintained for more than 24 months by repeated cloning and secretes IgM at concentrations of 2-8 μg/106cells/24h. The AC6C3 monoclonal antibody reacted with a cell surface component of ovarian tumor cell lines, as determined by cell surface immunofluorescence staining using the fluorescent activated cell sorter (FACS). In contrast, nylon wool nonadherent peripheral blood lymphocytes or red blood cells from normal donors were negative (<5% of the cells were stained). Immunoperoxidase staining with the AC6C3 monoclonal antibody of nonpermeabilized cryostat sections of freshly obtained or cryopreserved ovarian carcinoma specimens and human ovarian tumor xenografts demonstrated strong reactivity of these specimens. Most normal tissues Including brain, liver, heart, kidney and peritoneum demonstrated negative or weak reactions with AC6C3. Other carcinomas including breast, colon and some malignancies of neuroectodermal origin were strongly reactive with AC6C3. AC6C3 mediated complement-dependent cytotoxicity and identified a 32 Kd band in Western blotting and immunoprecipitation experiments conducted on surface labelled SKOV3 cells. The association constant for AC6C3 was determined at 2.3 X 1010M-1

ISSN:0272-457X    

DOI:10.1089/hyb.1991.10.21

年代:1991

数据来源: MAL

摘要:

The preparation of a panel of hybridoma cell lines secreting monoclonal antibodies (MAbs) specific for Recombinant Porcine Alpha I Interferon is reported. Of these MAbs, 28 were subcloned and 21 secreted their antibody during several months. They were partially characterized for their ability to bind and neutralize Alpha Interferons (IFNs-α) from different animal species. All the clones bound Recombinant Porcine IFN-α1, 8 bound Human leukocyte IFN, and one bound recombinant Human IFN-α2b, but not IFN-α2a. Two screening procedures were used for the detection of specific MAbs: neutralization assay and immunosorbent binding bioassay. This last method appears to be simple and very sensitive since it permits to detect 100pg/ml MAb. In addition it can detect weakly, as well as strongly neutralizing antibodies. Probable mechanisms involved in this assay, and possible applications of this method for IFNs-α subtyping are discu

ISSN:0272-457X    

DOI:10.1089/hyb.1991.10.35

年代:1991

数据来源: MAL

摘要:

The neutrophil (PMN) receptor for formylated peptides such as N-formyl-1-methionyl-1-leucyl-1-phenylalanine (FMLP) is involved in binding and subsequent response to certain chemotactic stimuli. The receptor on human PMN has been reported to consist of several glycoprotein components, ranging in size from 43-94 kDa. Furthermore, FMLP receptors on human PMN have been shown to contain both high and low affinity states. In this study, the receptor was purified by subjecting solubilized PMN plasma membrane components to FMLP-affinity chromatography, and was found to be comprised of four components, one of 68 kDa, and the others of 94, 48, and ≈40 kDa. Only the 68, the 94, and the ≈40 kDa components specifically bound a radioiodinated FMLP analogue. To further characterize these components, a battery of monoclonal antibodies reactive against the FMLP receptor was prepared. Seven monoclonal antibodies were selected on the basis of their reactivity with the 68 kDa receptor component. Some of these antibodies also cross-react with the 48 kDa component, suggesting that the 68 and the 48 kDa receptor moieties are immunologically related. These antibodies reacted with normal human neutrophils, but not with lymphocytes, or unstimulated HL-60 cells. Furthermore, the presence of 20 nmol of FMLP inhibited the binding of five of the anti-receptor antibodies to whole PMN. These results suggest that the epitopes recognized by these five antibodies may possibly be involved in FMLP bind

ISSN:0272-457X    

DOI:10.1089/hyb.1991.10.49

年代:1991

数据来源: MAL

摘要:

BALB/c mice were immunized with gelonin, a 30kD glycoprotein (type 1 RIP) from the seeds of Gelonium multiflorum. By polyethylene glycol-induced fusion of isolated spleen cells with the myeloma cell line NS-1, three different hybridomas were obtained. Two of them were found to secrete antibodies of the IgG1subclass, whereas the third cell line produced antibodies of the IgM type. The IgG1-secreting cell lines were adapted to serum-free medium conditions, and the antibodies were isolated from the culture supernatant. The isolated antibodies recognize independent epitopes on the gelonin molecule. The toxicity of gelonin in reticulocyte lysates was not affected when the protein was incubated with the antibodies. The IgG1s exhibit average affinity constants of about 109M-1and 1010M-1, respectively, as determined by a solid-phase EIA using the avidin-biotin system.

ISSN:0272-457X    

DOI:10.1089/hyb.1991.10.65

年代:1991

数据来源: MAL

摘要:

A monoclonal antibody (11G7) detecting a novel antigen on human hemopoietic progenitor cells (named 11G7R = 11G7 receptor) was raised by immunization of a Balb/c mouse with the leukemic blasts of a patient suffering from chronic myelogenous leukemia blast crisis (CML-BC). The antigen is expressed on most of MHC class II bearing peripheral blood leucocytes (PBL) and on a subpopulation of bone marrow mononuclear cells (BMMNC). By FACS-sorting and colony assays, it could be demonstrated that 11G7R is expressed on myelo-monocytic and myelo-granulocytic bone marrow precursor cells (GM-CFC, G-CFC, M-CFC) but is absent from erythroid precursor cells (BFU-E) and on cells exhibiting the capacity to form mixed colonies (GEMM-CFC). Double-fluorescence analysis on BMMNC revealed that 11G7R is expressed on a subset of B-cells, myeloid cells and cells carrying the HPCA-1 antigen (CD34). It has a similar distribution pattern to the myeloid antigens CD13 and CD33. However, in contrast to these antigens, 11G7R is also expressed on the blasts of several lymphoid leukemias (4/9 B-ALL, 1/2 T-ALL) and therefore it is not restricted to the myeloid lineage.

ISSN:0272-457X    

DOI:10.1089/hyb.1991.10.77

年代:1991

数据来源: MAL

摘要:

Five monoclonal antibodies (MAbs) were produced in a mouse hybridoma system against human placental glutathione transferase (GST п). Four of these monoclonal antibodies, named 461 to 464, were of immunoglobulin G class, whereas the monoclonal antibody 465 was of IgA class. All these MAbs specifically recognized the glutathione transferase from human placenta (class п ) showing no cross reactivity against the basic and the neutral forms of GST from human liver. When each MAb was incubated with the GST п, no inhibition of enzymatic activity towards 1-chloro-2,4-dinitrobenzene was observed except for MAb 465 which showed a slight inhibition to a serial dilution of 1:1

ISSN:0272-457X    

DOI:10.1089/hyb.1991.10.89

年代:1991

数据来源: MAL

摘要:

rasgenes have been shown to become oncogenes by single point mutations which result in amino acid substitutions that affect either their GTPase activity (positions 12,13,59,61) or their affinity for GTP and GDP.Rasoncogenes and their corresponding proteins have been described in a variety of human cancers as well as in animal tumors induced by physical and chemical carcinogens. One of these animal tumor systems involves the induction of mammary carcinomas in rats by a single dose of N-nitroso-N-methylurea (NMU), a methylating carcinogen. These NMU-induced mammary carcinomas contain transforming H-rasgenes activated by G -->A transitions in the second nucleotide of their 12th codon, presumably a consequence of the pre-mutagenic lesions induced by NMU. These G -->A mutations result in the replacement of the normal glycine in the 12th position of therasp21 protein by a glutamic acid residue. In this study, we report the generation of monoclonal antibodies (Mab) reactive with oncogenicrasp21 proteins containing glutamic acid at position 12 (p21 Glu-12). Mab designated E184 specifically recognized activatedrasp21 Glu-12 proteins but not normal p21 (Gly-12) or p21 proteins activated by other position 12 substitutions including arginine, aspartic acid, cysteine, valine or serine residues. Western blot analysis of NMU-induced mammary carcinomas demonstrated that Mab E184 recognized p21 proteins expressed in these rat tumors but not p21 present in normal tissues nor in other carcinogen-induced tumors known to carry H-rasoncogenes activated by mutations at position 61. Mab E184 should be a valuable reagent to determine the presence of activatedrasp21 (Glu-12) proteins in animal tumor systems and to correlate their expression with the initiation and/or progression of neoplastic development.

ISSN:0272-457X    

DOI:10.1089/hyb.1991.10.95

年代:1991

数据来源: MAL

摘要:

Monoclonal antibodies against a mouse preadipose cell line (MC3T3 -G2/PA6: PA6), which can support hemopoiesis by direct cell-to-cell interaction, were produced and characterized. The antibodies react with PA6 but not PA6-M (a mutant cell line) which has the capacity neither to contact with hemopoietic stem cells (HSCs) nor to support hemopoiesis. Endosteal cells in the bone marrow show positive staining to these antibodies. They inhibit pseudoemperipolesis of PA6 to HSCs, resulting in a significant decrease in hemopoietic cell number.These findings suggest that the monoclonal antibodies bind to the stromal cell receptors for HSCs and block the binding of HSCs to stromal cells leading to suppression of hemopoiesis.

ISSN:0272-457X    

DOI:10.1089/hyb.1991.10.103

年代:1991

数据来源: MAL