Hybridomas and Cellular Immortality
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Hybridomas and Cellular Immortality

Hybridomas and Cellular Immortality


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About the Book

The ability to "immortalize" immunologically-useful cells by hybridization with a unique cancer cell has revolutionized serological studies and has revealed new potential applications in all fields of biological sciences. This volume presents the studies from a highly successful national symposium on Hybridomas and Cellular Immortality held November 1981 in Houston, Texas. The individual chapters exhibit the diversity of topics discussed during the meeting. These include emphasis on the origin of antibody diversity, Band T lymphocyte differentiation, applications of monoclonal antibodies in studies of histocompatibility, tumor, and viral antigens, plus the use of somatic cell hybridizations for studying T cell products. Three papers focus on the emerging methodologies of in vitro primary immunizations for both humoral and cell-mediated immunities, relevant for coupling with hybridoma technology. There is a useful mix of general (methods) and specific (applications) chapters. A unique aspect of the book is the presentation of both recent research findings with concise descriptions of the state of the art methodologies. It is anticipated that this work will be of interest to a wide audience of practioners in biomedical research. Hopefully, the information contained will foster new and imagi- native ideas in hybridoma applications. Baldwin H. Tom, Ph.D. James P. Allison, Ph.D. vii CONTENTS PART L INTRODUCTION TO HYBRIDOMAS 1 Somatic Cell Hybrids and Hybridomas Baldwin H. Tom 3 1. Somatic Cell Hybrids 8 Hybridomas. * * * * * 2.

Table of Contents:
I. Introduction To Hybridomas.- 1 Somatic Cell Hybrids and Hybridomas.- 1. Somatic Cell Hybrids.- 2. Hybridomas.- References.- II. Antibody Diversity And Lymphocyte Differentiation.- 2 The Nature and Origin of Antibody Diversity.- 1. Introduction.- 2. V-region sequence diversity at the protein level.- 2.1. Nature of the sequence diversity.- 2.2. Origin of the sequence diversity.- 3. Analysis of Ig genes at the DNA level.- 3.1. General considerations, and the mouse X light chain Locus.- 3.2. The mouse ? light chain locus.- 3.3. Generation of VH-region diversity in mouse.- 3.4. Is additional V-region diversity introduced by somatic mutation?.- 4. Conclusions.- References.- 3 Hematopoietic Tumors: Normal or Abnormal Models of Leukocyte Differentiation?.- 1. Introduction.- 2. Cell surface phenotypes.- 2.1. Fc receptors.- 2.2. Thy-1.- 2.3. LEU-3.- 2.4. LY-1/LEU-1.- 2.5. E2.- 2.6. LEU-7.- 3. Discussion.- References.- 4 Dissection of Human Immunoregulatory T Lymphocytes: Implications for Understanding Clinical Disease.- 1. Summary.- 2. Introduction.- 3. Results and Discussion.- 3.1. Differentiation of T lymphocytes.- 3.2. Functions of mature T lymphocyte subsets.- 3.3. Clinical disorders of T lymphocytes.- 3.4. Human T cell malignant diseases.- 3.5. Immunotherapy by slective T cell subset manipulation.- 4. Experimental Procedures.- 4.1. Isolation of lymphoid populations.- 4.2. Production and characterization of monoclonal antibodies.- 4.3. Analysis and separation of lymphocyte populations with fluorescence activated cell sorter.- 4.4. Antigen stimulation of T lymphocytes.- 4.5. Detection of in vitro secretion of IgG.- References.- 5 Immune (?) Interferon and Lymphotoxin Production by T Cell Lines and Hybrids.- 1. Summary.- 2. Introduction.- 3. Results.- 3.1. Growth factor lines.- 3.1.1. Properties.- 3.1.2. Production of LT and IFN.- 3.2. T cell hybrids.- 3.2.1. Analysis.- 3.2.2. LT Production.- 4. Discussion.- 5. Experimental Procedures.- 5.1. T cell lines.- 5.2. T cell hybrids.- 5.3. Lymphotoxin assay.- 5.4. Interferon assay.- References.- III. Monoclonal Antibodies as Probes of Antigen Structure.- 6 Partial Structural Characterization of Human HLA-DR Subsets Using Monoclonal Antibodies.- 1. Summary.- 2. Introduction.- 3. Results and Discussion.- 3.1. Two monoclonal antibodies (L203 and L227) recognize subset of DR antigens.- 3.2. Both subsets of DR antigens are homolgous to murine I-E antigens.- 3.3. The antibody L227 recognizes isolated DR beta chains.- 3.4. Both monoclonal antibodies react with a third population of DR antigens.- 3.5. Amino acid sequence analysis of the L203 and L227 unique populations.- 3.6. The monoclonal antibodies IIIE3 and IVG1 recognize the same subset of DR antigens.- 3.7. III E3 recognizes a subset of L203 and L227 antigens.- 3.8. Amino acid sequence of the III E3/IVG1 antigens.- 4. Conclusions.- 5. Experimental Methods.- 5.1. Cells.- 5.2. Monoclonal antibodies and antisera.- 5.3. Isolation of radiolabeled Dr antigens.- 5.4. Lentil lectin affinity chromatography.- 5.5. Immunoprecipitation.- 5.6. Antibody affinity columns.- 5.7. SDS-Polycrylamide gel electrophoresis.- 5.8. Amino acid sequence determination.- References.- 7 Monoclonal Antibodies for Analysis of HLA Antigens: Further Studies with the W6/32 Antibody.- 1. Introduction.- 2. Results.- 2.1. Interaction of W6/32 antibody with HLA antigen.- 2.1.1. Binding of W6/32 to cells at equilibrium.- 2.1.2. Comparison of the binding of W6/32 IgG and FAB to LB cells at equilibrium.- 2.1.3. Kinetics of binding of W6/32 IgG and Fab to LB cells.- 2.1.4. Binding of W6/32 to soluble HLA antigens.- 2.1.5. Conclusions.- 2.2. Structural studies.- 2.2.1. Amino acid sequencing of W6/32 precipitates.- 2.2.2. Separation of exchange with an excess of non-radioactive "cold"-?-2 microglobulin...- 2.2.3. Separation of preparative SDS-PAGE.- References.- IV. Monoclonal Antibodies in Diagnosis and Therapy.- 8 Monoclonal Antibodies to Hepatitis B Surface Antigen.- 1. Summary.- 2. Introduction.- 3. Results.- 3.1. Fusions and characteristics of monoclonal anti-HBs.- 3.2. Immunoradiometric assay for HBsAg.- 4. Discussion.- 5. Experimental Procedures.- 5.1. Production of monoclonal antibodies to HBsAg.- 5.2. Characterization of monoclonal antibodies to HBsAg.- 5.3. Tissue culture.- 5.4. Assay procedure.- References.- 9 Wandering Around the Cell Surface: Monoclonal Antibodies Against Human Neuroblastoma and Leukemia Cell Surface Antigens.- 1. Summary.- 2. Introduction.- 3. Results.- 3.1. An anti-neuroblatoma antibody.- 3.2. The neuroblastoma determinant is also detectable on leukemia cells.- 3.3. Using a combination of monoclonal antibodies to detect neuroblastoma cells in marrow.- 3.4. Other monoclonal antibodies against neuroblastomas or leukemias.- 3.5. "Operationally tumor specific" antibodies.- 3.6. In vitro immunization.- 3.7. Detection of antibodies inhibiting leukemia cell proliferation.- 3.8. Isolation of cell surface molecules using monoclonal antibodies.- 3.9. Monoclonal antibodies can be used to isolate minor components of the cell surface.- 3.10. Concluding summary.- 4. Discussion.- 5. Experimental Procedures.- 5.1. Production and characterization of monoclonal antibodies.- 5.2. In vitro immunization.- 5.3. Detection of monoclonal antibodies inhibiting Reh cell proliferation.- 5.4. Affinity chromatography of neuroblastoma antigens.- References.- 10 Monoclonal Antibodies to Human Melanoma-Associated Antigen p97.- 1. Summary.- 2. Introduction.- 3. Results.- 3.1. Monoclonal antibodies to p97.- 3.2. Tissue distribution of p97.- 3.3. Structural studies of p97.- 3.4. Relationship of p97 to other tumor-associated antigens.- 4. Discussion.- 4.1. Screening hybridomas.- 4.2. Antigen specificity.- 4.3. Comparison of tumor-associated antigens.- 4.4. Nature, function, and practical application of tumor-associated antigens defined by monoclonal antibodies.- References.- 11 A Biochemical and Biosynthetic Analysis of Human Melanoma-Associated Antigens with Monoclonal Antibodies.- 1. Summary.- 2. Introduction.- 3. Results.- 4. Discussion.- 5. Materials and Methods.- 5.1. Cells.- 5.2. Monoclonal antibodies.- 5.3. Serological assays.- 5.4. Biosynthetic labelling procedures.- 5.5. Indirect immunoprecipitation/SDS-PAGE.- 5.6. Cell fractionation procedure.- 5.7. Materials.- References.- 12 Monoclonal Antibodies to a Tumor Specific Antigen on Rat Mammary Carcinoma Sp4 and their use in Drug Delivery Systems.- 1. Introduction.- 2. Results and Discussion.- 2.1. Monoclonal antibody specifying tumor specific antigen associated with rat mammary carcinoma Sp4.- 2.2. Anti-tumor activities of Sp4/A4 monoclonal antibody.- 2.3. In vivo localization of Sp4/A4 monoclonal antibody.- 2.4. Anti-tumor activity of Sp4/A4 antibody-adriamycin conjugates.- 3. Conclusions.- 4. Materials and Methods.- 4.1. Tumors.- 4.2. Mammary carcinoma Sp4 monoclonal antibody (Sp4/A4).- 4.3. Treatment of Sp4 with Sp4/A4 monoclonal antibody.- 4.4 Complement-dependent cytotoxicity test.- 4.5. Antibody-dependent cell-mediated cytotoxicity.- 4.6. Radiolabelling of Sp4/A4 monoclonal antibody for in vivo localization.- 4.7. Antitumor activity of Sp4/A4 monoclonal antibody conjugated to adriamycin.- References.- 13 Monoclonal Antibody to a Tumor Specific Epitope of Murine Lymphoma Cells: Use in Characterization of Antigen and in Immunotherapy.- 1. Summary.- 2. Introduction.- 3. Results and Discussion.- 3.1. Production of monoclonal antibody.- 3.2. Reactivity of MAb 124-40 with C6XL lymphoma cells.- 3.3. Lack of reactivity of MAb 124-40 with other normal or malignant lymphoid cells.- 3.4. Characterization of Ag 124-40.- 3.5. Expression of similar cell surface component by normal T cells and thymocytes.- 3.6. Immunotherapy with MAb 124-40 and pokeweed antiviral protein conjugate.- 4. Conclusions.- 5. Experimental Procedures.- 5.1. Mice.- 5.2. Cell Lines.- 5.3. Hybridoma construction.- 5.4. Antibody binding assay.- 5.5. Flow microfluorometry.- 5.6. Radioiodination.- 5.7. Immunoprecipitation.- 5.8. Polyacrylamide gel electrophoresis.- References.- V. New and Relevant Methodology.- 14 Human Hybridomas.- 1. Summary.- 2. Introduction.- 3. Results.- 4. Discussion.- 5. Experimental Methods.- 5.1. Selection of somatic cell hybrids.- 5.2. Production of human immunoglobulin (Ig) chain.- 5.3. Isozyme analysis.- 5.4. Chromosome analysis.- References.- 15 Procedures for In Vitro Immunization and Monoclonal Antibody Production.- 1. Summary.- 2. Introduction.- 3. Results.- 4. Discussion.- 5. Methods.- 5.1. Culture conditions.- 5.2. Immunization in culture.- 5.3. Hybridization.- 5.4. Enzyme linked immunosorbent assay (ELISA).- 5.5. HPLC-ELISA.- 5.6. Immunogold preparation.- 5.7. Immunogold staining.- 5.8. Immunogold separation.- References.- 16 Liposome-facilitated In Vitro Induction of Primary Cell-Mediated Immunity to Human Cancer Antigens: Potential Adjunct to Hybridoma Technology.- 1. Summary.- 2. Introduction.- 2.1. In vitro immunization.- 2.2. cell-free immunizations.- 2.3. Liposome-mediated immunity.- 3. Results.- 3.1. Summary of studies.- 3.2. In vitro xenoimmunization to human tumor antigens.- 3.3. In vitro alloimmunization to human tumor antigens.- 4. Discussion and Conclusions.- 5. Experimental Procedures.- 5.1. Preparation of tissue culture cells.- 5.2. Preparation of tumor cell membranes.- 5.3. Preparation of liposome-membrane antigens.- 5.4. Quantitation of protein antigens.- 5.5. Quantitation of liposomes.- 5.6. In vitro immunization.- 5.6.1. Generation of cytotoxic activity with liposomes.- 5.6.2. Inhibition of cytotoxicity.- 5.6.3. Cell-mediated cytotoxicity assay.- 5.6.4. Elicitation of blastogenic response.- References.- 17 Comparison of Three Immunoassays for Screening Anti-Hepatitis B Hybridomas.- 1. Summary.- 2. Introduction.- 3. Results and Discussion.- 4. Experimental Procedures.- 4.1. Antisera.- 4.2. Antibody-enzyme conjugation.- 4.3. Immunoassays.- References.- VI. Poster Abstracts.- 1. T-Lymptocyte Subsets of Rhesus Macaque as Determined by Monoclonal Antibodies: T?, T?, T-NULL, and Histamine Receptor Bearing T Lymphocytes.- 2. Production of Human Immunoglobulin Secreting Clones Without the Use of H.A.T. Sensitive Fusion Partners.- 3. Species Distribution of Antigenic Sites on Mouse LDH-C4 Determined by Monoclonal Antibodies.- 4. A New Non-Immunoglobulin Producing Human Myeloma Cell line for Production of Hybridomas.- 5. Monoclonal Antibodies to Human Prostate and Bladder Tumor Associated Antigens.- 6. Antibody Purification by Recycling Isoelectric Focusing.- 7. Monoclonal Antibodies Against Human Ejaculated Sperm-Application in Antibody Mediated Infertility.- 8. Analysis of Peripheral Blood and Salivary Gland Lymphocytes in Sjogren's Syndrome.- 9. A Monoclonal Antibody Which Identifies a Subpopulation of Human B Lymphocytes and Monocytes.- 10. In Vivo Localization of Tumor Associated Antigens with Radioactively Labeled Monoclonal Antibodies.- 11. Characteristics of Monoclonal Antibodies to Herpes Simplex Type 2.- 12. Type Specific Monoclonal Antibodies to Equine Infectious Anemia Virus: Development and Use in the Characterization of Canine and Feline Derived Viruses.- 13. Biochemical and Biological Analysis of Venezuelan Equine Encephalomyelitis Viruses (VEE) Using Monoclonal Antibodies.- 14. Monoclonal Antibodies Specific for Herpes Simplex Virus Type 1 Mediate Antiviral Effects In Vitro and In Vivo.- 15. Monoclonal Antibodies Against Antigens of Amastigotes and Epimastigotes of Trypanosma Cruzi.- 16. Differential Effects of Four Monoclonal Antibodies to Human ?2-Microglobulin on In Vitro Lymphocyte Functions.- 17. Selective Inhibition of Functional Sites on Cell Bound Human C3b By Monoclonal Antibodies.- 18. Generation of Monoclonal Antibodies Reacting With Human Normal and Neoplastic Mammary Duct Epithelial Cells.- 19. Resolution of Human Liver Mao A and B Using A Monoclonal Antibody Immunoaffinity Column.- 20. Isolation of Antibodies that are Specific for Human Carboxypeptidase B But Not for the Zymogen Procarboxypeptidase B by Affinity Chromatography.- 21. Dexamethasone Effects on Hybridoma Formation.- 22. Class Antigen in the Rat: Evidence for More Than One Using Monoclonal Antibodies.- 23. An Immunohistochemical Method for Large Scale Specificity Screening of Monoclonal Antibodies.- 24. Monoclonal Antibodies to Chicken Hemopoietic-Lymphoid Cellular Differentiation Antigens.- Author Index.


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Product Details
  • ISBN-13: 9780306414671
  • Publisher: Springer Science+Business Media
  • Publisher Imprint: Kluwer Academic/Plenum Publishers
  • Language: English
  • ISBN-10: 0306414678
  • Publisher Date: 01 Oct 1983
  • Binding: Hardback
  • Returnable: N


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