Production of Biologicals from Animal Cells in Culture

Production of Biologicals from Animal Cells in Culture reviews the state of the art in animal cell biotechnology, with emphasis on the sequence of events that occur when generating a biological from animal cells in culture. Methods that enable adjustment of nutrient feed streams into perfusion bioreactors so as to increase productivity are described. A number of issues are also addressed, such as the usefulness of the fingerprint method for cell characterization.

Comprised of 135 chapters, this book begins with an overview of the problems and benefits of animal cell culture, followed by a discussion on the isolation of immortal murine macrophage cell lines. The reader is systematically introduced to the use of DNA fingerprinting to characterize cell banks; immortalization of cells with oncogenes; lipid metabolism of animal cells in culture; and energetics of glutaminolysis. Subsequent chapters explore serum-free and protein-free media; the physiology of animal cells; gene expression in animal cell systems; and animal cell bioreactors. The monitoring and assay of animal cell parameters are also considered, along with downstream processing and regulatory issues.

This monograph will be of interest to students, practitioners, and investigators in the fields of microbiology and biotechnology.

Hyclone Award Lecture

1. Animal Cell Culture: The Problems and Rewards

Section 1 Cell Lines and their Characterization

2. The Isolation of Immortal Murine Macrophage Cell Lines

3. Propagation of Human CD4-Immunoglobulin Producing Cell Lines

4. Characterization of Cell Banks by DNA Fingerprinting

5. Evaluating the Safety of Murine and Human Hybridomas: New Problems and New Techniques

6. Defective Endogenous Retroviruslike Particles of Chinese Hamster Ovary Cells

7. Reference Cell Lines

8. A European Database for Animal Cell Lines

9. Cell Line Data Base: an On-Line Data Bank Where Industry Meets Researchers

10. Adhesion and Motility of Embryonic and Cancer Cells

11. Immortalization of Cells with Oncogenes

Section 2 Nutrient Media with Special Supplements

12. Lipid Metabolism of Animal Cells in Culture - A Review

13. Lipoprotein Requirements of Cultured Mammalian Cells

14. Effect of Nutrients and Waste Metabolites on Growth, Metabolism and Protein Synthesis in Hybridoma

15. Hybridoma Growth, Metabolism, and Product Formation in Hepes-Buffered Medium: Effect of Passage Number and pH

16. A Screening Experiment of Media Supplements on CHO Cells

17. Energetics of Glutaminolysis - A Theoretical Evaluation

18. Degrading Effect of Light on Cell Culture Media

19. Culture of Hybridoma in Dialysed Media

20. Evaluation of Fetal Bovine Serum Substitutes on Cell Growth and Recombinant Protein Production

21. Cell Growth Activators (CGA) and Cell Function Modulators (CFM) of Blood: Effect on Cell Replication and Protein Expression

22. Increased Expression of Factor VIII by Butyrate in Chinese Hamster Ovary Cells

23. Polyamine Enhanced Product Expression from Transformed and Recombinant Cell Lines

24. The Development of a Process for the Production of HIV1 GP120 from Recombinant Cell Lines

25. 'Safe' Animal Derived Medium Supplements for Cell Cultures

26. Effect of Desferrioxamine on Transferrin Receptor Expression, Growth and IgG Production of the Rat Hybridoma MA C57

Section 3 Serum-Free and Protein-Free Media

27. Development of Serum-Free Media for Mammalian Cell Culture

28. Concept and Strategy for the Development of a General Purpose Serum-Free Medium for Anchorage-Dependent Cells

29. Growth of Mammalian Cell Lines Under Serum-Free Conditions

30. Development of a Low-Serum Growth Medium for Production of a Human Glycoprotein by Recombinant BHK Cells

31. Optimalisation of a Serum Free Medium for Large Scale Continuous Growth and Immunoglobulin Production by a Mouse Hybridoma

32. Longterm Serum-Free Culture of Murine Hybridomas

33. Serum-Free Media Suitable for Upstream and Downstream Processing

34. Production of Human Fibronectin from Serum-Free Continuous Cultured Human Hepatoblastoma Line HUH-6

35. Development of a Protein-Free Medium for Bioreactor Cultures of Hybridoma Cell Lines

36. Conditions for the Production of Recombinant IL-2 in Stirred Suspension Culture Using a Protein-Free Medium

37. A Protein-Free Serum Replacement (SSR) for Cell Culture

38. Adaptation and Characterization of Hybridomas Growing in Low Protein and Protein-Free Medium

39. Characterization of Protease Activity in Serum-Free Culture Supernatants of Hybridomas and Recombinant Mammalian Cells

Section 4 Cell Physiology

40. Animal Cells in Culture Make Secondary Metabolites too

41. Removal of Inhibitory Factors from Hybridoma Cell Cultures by Gel Filtration

42. The Effect of Metabolic by-Products on Animal Cells in Culture

43. Cytoskeletal Microfilament Network and Energy Metabolism Affect Ability of Animal Cells to Resist Shear Injury

44. Intracellular Concentration of ATP and other Nucleotides During Continuous Cultivation of Hybridoma Cells

45. Determination of the 'Critical Shear Stress Level' for Adherent BHK Cells

46. Effects of Dissolved Oxygen Supply and Glucose Concentration on Human Lymphoblastoid Cell Growth and Metabolism in Continuous Culture

47. The Role of Dissolved Oxygen in Growth of Animal Cells

48. The Effect of Osmolarity on Hybridoma Cell Growth and Antibody Production in Serum-Free Growth Media

49. Physiological Investigations in High Density Perfusion Culture of Free Suspended Animal Cells

50. Factors Affecting Hybridoma Cell Attachment to Derivatized Polyacrylamide Beads

51. Adaption of Anchorage-Dependent Cells to Glutamine-Free Medium

52. The 2'5' Oligoadenylate Synthetase, a Multienzyme System Involved in the Growth Properties and the Sensitivity to Viruses of Mammalian Cells: Selection of Antibodies in Order to Investigate their Specific Biological Roles

Section 5 Gene Expression in Animal Cell Systems

53. Recent Developments in Mammalian Expression Systems

54. Genetic Engineering of Cellular Physiology

55. Stability of Amplified DNA in Chinese Hamster Ovary Cells

56. Methotrexate and CHO Cells: Productivity and Genetics of Amplified Expression Vector Sequences

57. The Stability of Expression of Hybridoma Cells in Homogeneous Continuous Culture Systems

58. Cloning and Expression of a Bovine Anti-Testosterone Monoclonal Antibody

59. Use of Dicistronic Transcription Units for the Correlated Expression of Two Genes in Mammalian Cells

60. Large-Scale Recombinant Protein Production Using the Insect Cell-Baculovirus Expression System: Antistatin and β-Adrenergic Receptor

61. Quantitation of mRNA Specific for Heavy and Light Chains of IgG in Hybridomas During Different Phases of Batch Culture

Section 6.1 Bioreactors: Overview

62. Development of Mammalian Cell Manufacturing Processes

63. A New Look at Animal Cell Bioreactor Development

64. Mammalian Cell Fermentation: Are Specialized Bioreactors a Necessity?

Section 6.2 Bioreactors: Hardware

65. Engineering for Sterility and Containment: Design of a 225 L 'Polymodal' Fermenter for the Containment of P3 Pathogens

66. Monoclonal Antibody Production Using an Airlift Fermentation System Consisting of a Continuous Seed Fermenter and a Fed Batch Production Fermenter

67. Bleeding Out Dead Cells from Fermentation Systems

68. High Intensity Growth of Adherent Cells on a Porous Ceramic Matrix

69. Considerations in the Design, Development and Scale-Up of Glass Bead Packed Reactors

70. On-Line Removal of Cells from Continuous Suspension Cultures

Section 6.3 Bioreactors: Particles

71. Microsphere-Induced Aggregate Culture of Animal Cells

72. The Growth of CHO and BHK Cells as Suspended Aggregates in Serum-Free Medium

73. CultiSpher - Macroporous Gelatin Microcarrier - New Applications

74. Cultivation of CHO cells on Microcarriers Dormaceli

Section 6.4 Bioreactors: Membranes and Perfusion

75. Fermentation of AT III Producing BHK Cells in a Double Membrane Perfusion Bioreactor

76. Oxygen Transfer Characteristics in Cell Culture Fermenters: Direct Sparging-Membrane Oxygenation-Bubble-Free Aeration Through a Rotating Sieve

77. Hybridoma Culture in a Membrane System: The Influence of Oxygen on a Reactor's Performance and on the Evaluation of Cellular Density

78. Hybridoma, Antibody and Growth Factor Distributions in the Shell-Side of Ultrafiltration Hollow Fiber Bioreactors

79. Cultivation of Insect Cell Lines in Stirred Membrane Reactors

80. High Density Culture of Hybridoma Cells Using a Perfusion Culture Apparatus with Multi-Settling Zones

81. High Cell Density Perfusion Culutre of Insect Cells for Production of Baculovirus and Recombinant Protein

Section 6.5 Bioreactors: Optimization via Metabolism

82. Metabolite Parameters Modulating Monoclonal Antibody Production by Hybridoma Cells in Flask and Macroporous Glass Sphere Culture

83. Optimization of OKT3 Hybridoma Cultures in a Perfusion Stirred Reactor

84. Biochemical Control of Monoclonal Antibody Secretion in Hollow Fibre Reactors

85. Optimization of Human Anti-Rhesus IgG Production Using Hollow Fiber Technology

86. Parameters for Optimization of Fermentation of an AT III Producing Cell Line

87. Enhanced Productivity of Hybridoma and Recombinant CHO Cell Cultures by Pluronic F-68 and other Medium Components, and by Increased Perfusion Rates, in Fluidized-Bed Bioreactors

88. Improved Performance of the Fluidized Bed Reactor for the Cultivation of Animal Cells

Section 6.6 Bioreactors: Comparative Studies

89. Bioreactors for Production of HIV - A Comparative Evaluation of Flask, Suspension and Fixed Bed Cultures

90. Comparison of Semicontinuous Culture and Perfusion Culture in a 75 L Bioreactor

91. Comparison of Two Systems Based on Porous Beads for the Production of Recombinant von Willebrand Factor by CHO Cells

92. Comparison of the Production Efficiency of Mammalian Cells Grown in a Fluidized Bed and in a Stirred Tank Reactor

93. Monoclonal Antibody Production of Three Different Hollow Fibre Bioreactor Configurations

94. A Comparison of Two Reactors for Cell Cultivation

Section 7 Monitoring and Assay of Animal Cell Parameters

95. Staining of Chinese Hamster Ovary Cells on Macroporous Matrices

96. The Role of Oxidation-Reduction Potential in Monitoring Growth of Cultured Mammalian Cells

97. Potential and Pitfalls of Using LDH Release for the Evaluation of Animal Cell Death Kinetics

98. Flow Cytometric (FCM) Analysis on the Genetic Stability of Continuous Cell Lines

99. Biomass Estimation in Anchorage-Dependent Animal Cell Cultures

100. Dynamic Assay of Synthetic Activities in Single Hybridoma Cells

101. Rapid Monitoring of Monoclonal Antibodies in Cell Culture Media by High Performance Liquid Affinity Chromatography (HPLAC)

102. Application of the PCFIA to Rapid Off-Line Process Monitoring

Section 8 Kinetics and Modeling

103. Mathematical Estimator for the Evaluation of Cell Density and Medium Composition in Hybridoma Cultures

104. Kinetic Effect of Growth Factors on Batch and Continuous Hybridoma Cultures

105. Batch Production Kinetics of Hybridomas: Pulse Experiments

106. Analysis of Hybridoma Growth, Metabolism and Product Formation in Continuous Suspension Culture

107. Kinetics of Nutrient Utilization in a Hybridoma Batch Culture

108. Modeling Growth of and Antibody Production by Hybridomas in Glutamine Limited Suspension Cultures

109. A Computer Simulation of the Kinetics and Energetics of Hybridoma Cell Growth and Antibody Production

110. A Kinetic Model for the Influence of Serum in Batch and Continuous Hybridoma Cultures

Section 9 Downstream Processing

111. Isolation of Biologicals: Leachables and Cleaning in Place Become a Crucial Issue in Liquid Chromatography

112. Composite High Flow Rate Ion Exchangers for Economical Scale-Up

113. Serum Free Media (SFM) and Serum Substitutes (SS) in Animal Cell Culture: Downstream Considerations

114. The Use of Affinity Chromatography for the Purification of Monoclonal Antibodies

Section 10 Products

115. Production of a Pharmaceutical Enzyme: Animal Cells or E. coli?

116. Oral Polio Vaccine (Sabin) Produced on Large-Scale Vero Cell Culture

117. Strategies for Improved Production of Epstein-Barr Virus

118. Optimal Conditions for the Growth of Pig Kidney Endothelial Cells (SK) Proliferated on Microcarriers and Production of Pig Parvovirus (PPV) Vaccine

119. Continuous Production of a Novel Thrombolytic from 50/100 Litre Perfused Microcarrier Cultures

120. Production of Recombinant Human Thyrotropin and its Incomplete Processing

121. Production of a Monoclonal Antibody Against Gelsolin

122. Appearance of Nonspecific Amounts of Monoclonal Antibody During Fermentation Caused by Decreased Cell Viability

123. Production of Biologically Active Insulin-Like Growth Factor by Cells Transfected with a Recombinant Retroviral Vector Encoding IGF-1 726

124. In vitro Production of Growth Regulators for Human Epithelial Cells

125. Purification and Characterization of Immunoglobulin Production Stimulating Factor II? Derived from Namalwa Cells

126. Production of a Chimeric Antibody from Tumor Imaging and Therapy from Chinese Hamster Ovary (CHO) and Myeloma Cells

127. Optimization of tPA Production in a 3L Continuous Perfusion Bioreactor

128. The Production and Assay of tPA from Mammalian Cells

129. Constitutive Secretion of Soluble CD23 Receptors in Hollow Fiber Reactors

130. Production of Proteins from Cell Nuclei by Cyclic Continuous Fermentation

Section 11 Regulatory Issues

131. Biologicals and Regulatory Aspects in the EEC

132. Mapping of N-Linked Oligosaccharides from Human Recombinant Glycopro Teins: Quality Control of Glycosylated Therapeutic Polypeptides

133. Validation of Downstream Processing

134. The Culture Environment Affects Recombinant Protein Processing in CHO Cells

135. The Medi-Cult Hybritest - A New Test for in Vitro Toxicology

List of Exhibitors

List of Participants

Subject Index

Author Index