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Culture of Animal Cells : A Manual of Basic Technique and Specialized Applications.

By: Material type: TextTextSeries: New York Academy of Sciences SeriesPublisher: Newark : John Wiley & Sons, Incorporated, 2016Copyright date: ©2016Edition: 1st edDescription: 1 online resource (758 pages)Content type:
  • text
Media type:
  • computer
Carrier type:
  • online resource
ISBN:
  • 9781118873373
Subject(s): Genre/Form: Additional physical formats: Print version:: Culture of Animal CellsDDC classification:
  • 571.6/381
LOC classification:
  • QH585.2 .F747 2016
Online resources:
Contents:
Intro -- CULTURE OF ANIMAL CELLS -- Contents -- List of Figures -- List of Color Plates -- List of Tables -- List of Protocols -- List of Minireviews -- Preface and Acknlowledgments -- Abbreviations -- About the Companion Website -- 1. Introduction -- 1.1 Historical Background -- 1.2 Advantages of Tissue Culture -- 1.2.1 Control of the Environment -- 1.2.2 Characterization and Homogeneity of Sample -- 1.2.3 Economy, Scale, and Mechanization -- 1.2.4 In vitro Modeling of In vivo Conditions -- 1.3 Limitations -- 1.3.1 Expertise -- 1.3.2 Quantity -- 1.3.3 Dedifferentiation and Selection -- 1.3.4 Origin of Cells -- 1.3.5 Instability -- 1.4 Major Differences In Vitro -- 1.5 Types of Tissue Culture -- References -- 2. Biology of Cultured Cells -- 2.1 The Culture Environment -- 2.2 Cell Adhesion -- 2.2.1 Cell Adhesion Molecules -- 2.2.2 Intercellular Junctions -- 2.2.3 Cytoskeleton -- 2.2.4 Extracellular Matrix -- 2.2.5 Cell Motility -- 2.3 Cell Proliferation -- 2.3.1 Cell Cycle -- 2.3.2 Control of Cell Proliferation -- 2.4 Differentiation -- 2.4.1 Induction and Maintenance of Differentiation -- 2.4.2 Plasticity of Differentiation and Dedifferentiation -- 2.5 Cell Signaling -- 2.6 Energy Metabolism -- 2.7 Origin of Cultured Cells -- 2.7.1 Initiation of the Culture -- 2.7.2 Evolution of Cell Lines -- 2.7.3 Senescence -- 2.7.4 Transformation and the Development of Continuous Cell Lines -- 2.8 Definitions -- References -- Supplier -- 3. Laboratory Design and Layout -- 3.1 Planning, Furnishing, and Services -- 3.1.1 Requirements -- 3.1.2 Services -- 3.1.3 Ventilation -- 3.2 Design and Layout -- 3.2.1 Sterile Handling Area -- 3.2.2 Laminar Flow Hoods -- 3.2.3 Service Bench -- 3.2.4 Quarantine and Containment -- 3.2.5 Incubation -- 3.2.6 Preparation Area -- 3.2.7 Storage -- 3.3 Disaster Management -- References -- 4. Equipment and Materials.
4.1 Requirements of a Tissue Culture Laboratory -- 4.2 Sterile Working Area -- 4.2.1 Laminar-Flow BSC -- 4.2.2 Service Carts -- 4.2.3 Sterile Liquid Handling-Pipetting and Dispensing -- 4.2.4 Inverted Microscope -- 4.2.5 Camera and Monitor -- 4.2.6 Dissecting Microscope -- 4.2.7 Centrifuge -- 4.2.8 Cell Counting -- 4.3 Incubation and Culture -- 4.3.1 Incubator -- 4.3.2 Humid CO2 Incubator -- 4.3.3 Temperature Recorders -- 4.3.4 Roller Racks -- 4.3.5 Magnetic Stirrer -- 4.3.6 Culture Vessels -- 4.4 Preparation and Sterilization -- 4.4.1 Washup -- 4.4.2 Preparation of Media and Reagents -- 4.4.3 Sterilization -- 4.5 Storage -- 4.5.1 Consumables -- 4.5.2 Refrigerators and Freezers -- 4.5.3 Cryostorage Containers -- 4.5.4 Controlled-Rate Freezer -- 4.6 Supplementary Laboratory Equipment -- 4.6.1 Computers and Networks -- 4.6.2 Upright Microscope -- 4.6.3 Low-Temperature Freezer -- 4.6.4 Confocal Microscope -- 4.6.5 PCR Thermal Cycler -- 4.7 Specialized Equipment -- 4.7.1 Microinjection Facilities -- 4.7.2 Colony Counter -- 4.7.3 Centrifugal Elutriator -- 4.7.4 Flow Cytometer -- References -- Suppliers -- 5. Aseptic Technique -- 5.1 Objectives of Aseptic Technique -- 5.1.1 Risk of Contamination -- 5.1.2 Maintaining Sterility -- 5.2 Elements of Aseptic Environment -- 5.2.1 Laminar Flow -- 5.2.2 Quiet Area -- 5.2.3 Work Surface -- 5.2.4 Personal Hygiene -- 5.2.5 Reagents and Media -- 5.2.6 Cultures -- 5.3 Sterile Handling -- 5.3.1 Swabbing -- 5.3.2 Capping -- 5.3.3 Flaming -- 5.3.4 Handling Bottles and Flasks -- 5.3.5 Pipetting -- 5.3.6 Large-Volume Dispensing -- 5.3.7 Pouring -- 5.4 Standard Procedure -- Protocol 5.1. Aseptic Technique In BSC -- Protocol 5.2. Working On The Open Bench -- 5.4.1 Culture Flasks and Bottles -- 5.4.2 Petri Dishes and Multiwell Plates -- Protocol 5.3. Handling Dishes or Plates -- 5.5 Apparatus and Equipment.
5.5.1 Refrigerators and Coldrooms -- 5.5.2 Incubators -- Protocol 5.4. Cleaning Incubators -- 5.5.3 Boxed Cultures -- 5.5.4 Gassing with CO2 -- References -- Suppliers -- 6. Safety, Bioethics, and Validation -- 6.1 Laboratory Safety -- 6.2 Risk Assessment -- 6.3 Standard Operating Procedures -- 6.4 Safety Regulations -- 6.5 General Safety -- 6.5.1 Operator -- 6.5.2 Equipment -- 6.5.3 Glassware and Sharp Items -- 6.5.4 Chemical Toxicity -- 6.5.5 Gases -- 6.5.6 Liquid Nitrogen -- 6.5.7 Burns -- 6.6 Fire -- 6.7 Ionizing Radiation -- 6.7.1 Ingestion -- 6.7.2 Disposal of Radioactive Waste -- 6.7.3 Irradiation from Labeled Reagents -- 6.7.4 Irradiation from High-Energy Sources -- 6.8 Biohazards -- 6.8.1 Levels of Biological Containment -- 6.8.2 Biological Safety Cabinets (BSCs) -- 6.8.3 Human Biopsy Material -- 6.8.4 Cell Lines -- 6.8.6 Genetic Manipulation -- 6.8.6 Disposal of Biohazardous Waste -- 6.8.7 Decontamination and Fumigation -- 6.9 Bioethics -- 6.9.1 Animal Tissue -- 6.9.2 Human Tissue -- 6.10 Quality Assurance -- 6.10.1 Procedures -- 6.10.2 Quality Control (QC) -- 6.10.3 Validation -- 6.10.4 Authentication -- 6.10.5 Provenance -- 6.10.6 Contamination -- References -- Suppliers -- 7. Culture Vessels and Substrates -- 7.1 The Substrate -- 7.1.1 Attachment and Growth -- 7.1.2 Common Substrate Materials -- 7.1.3 Alternative Substrates -- 7.2 Treated Surfaces -- 7.2.1 Substrate Coating -- Protocol 7.1. Preparation of ECM -- 7.2.2 Feeder Layers -- 7.2.3 Nonadhesive Substrates -- 7.3 Choice of Culture Vessel -- 7.3.1 Cell Yield -- 7.3.2 Multiwell Plates -- 7.3.3 Flasks and Petri Dishes -- 7.3.4 High Yields -- 7.3.5 Suspension Culture -- 7.3.6 Venting -- 7.3.6 Sampling and Analysis -- 7.3.7 Uneven Growth -- 7.3.8 Cost -- 7.4 Specialized Systems -- 7.4.1 Permeable Supports -- 7.4.2 Three-Dimensional Matrices -- References -- Suppliers.
8. Defined Media and Supplements -- 8.1 D evelopment of Media -- 8.2 Physicochemical Properties -- 8.2.1 pH -- Protocol 8.1. Preparation of pH Standards -- 8.2.2 CO2 and Bicarbonate -- 8.2.3 Buffering -- 8.2.4 Oxygen -- Minireview 8.1. Hypoxic Cell Culture -- 8.2.5 Osmolality -- 8.2.6 Temperature -- 8.2.7 Viscosity -- 8.2.8 Surface Tension and Foaming -- 8.3 Balanced Salt Solutions -- 8.4 Complete Media -- 8.4.1 Amino Acids -- 8.4.2 Vitamins -- 8.4.3 Salts -- 8.4.4 Glucose -- 8.4.5 Organic Supplements -- 8.4.6 Hormones and Growth Factors -- 8.4.7 Antibiotics -- 8.5 Serum -- 8.5.1 Protein -- 8.5.2 Growth Factors -- 8.5.3 Hormones -- 8.5.4 Nutrients and Metabolites -- 8.5.5 Lipids -- 8.5.6 Minerals -- 8.5.7 Inhibitors -- 8.6 Selection of Medium and Serum -- 8.6.1 Serum Batch Reservation -- 8.6.2 Testing Serum -- 8.6.3 Heat Inactivation -- 8.7 Other Supplements -- 8.7.1 Amino Acid Hydrolysates -- 8.7.2 Embryo Extract -- 8.7.3 Conditioned Medium -- 8.8 Storage -- References -- Suppliers -- 9. Serum-Free Media -- 9.1 Disadvantages of Serum -- 9.2 Advantages of Serum-Free Media -- 9.3 Disadvantages of Serum-Free Media -- 9.4 Replacement of Serum -- 9.4.1 Commercially Available Serum-Free Media -- 9.4.2 Serum Replacements -- 9.4.3 Serum-Free Subculture -- 9.4.4 Hormones -- 9.4.5 Growth Factors -- 9.4.6 Nutrients in Serum -- 9.4.7 Proteins and Polyamines -- 9.4.8 Viscosity -- 9.5 D evelopment of Serum-Free Medium -- 9.6 Selection of Serum-Free Medium -- 9.6.1 Cell or Product Specificity -- 9.6.2 Adaptation of Cell Lines to Serum-Free Media -- 9.7 Preparation of Serum-Free Medium -- 9.8 Animal Protein-Free Media -- 9.9 Conclusions -- References -- Suppliers -- 10. Preparation and Sterilization -- 10.1 Preparation of Reagents and Materials -- 10.2 Sterilization of Apparatus and Liquids -- 10.2.1 Dry-Heat Sterilization -- 10.2.2 Pressurized-Steam Sterilization.
10.2.3 Irradiation -- 10.2.4 Chemical Sterilization -- 10.2.5 Sterility Indicators -- 10.2.6 Filter Sterilization -- 10.3 Apparatus -- 10.3.1 Glassware -- Protocol 10.1. Preparation and Sterilization of Glassware -- 10.3.2 Glass Pipettes -- Protocol 10.2. Preparation and Sterilization of Glass Pipettes -- 10.3.3 Screw Caps -- Protocol 10.3. Preparation and Sterilization of Screw Caps -- 10.3.4 Selection of Detergent -- 10.3.5 Miscellaneous Equipment -- 10.3.6 Reusable Sterilizing Filters -- Protocol 10.4. Sterilizing Filter Assemblies -- 10.4 Reagents and Media -- 10.4.1 Water -- Protocol 10.5. Preparation and Sterilization of Ultrapure Water (UPW) -- 10.4.2 Maintenance of Water Purifier -- 10.4.3 Balanced Salt Solutions -- Protocol 10.6. Preparation and Sterilization of D-PBSA -- 10.4.4 Preparation and Sterilization of Media -- Protocol 10.7. Preparation of Medium from 1× Stock -- Protocol 10.8. Preparation of Medium from 10× Concentrate -- Protocol 10.9. Preparation of Medium from Powder -- 10.5 Sterilization of Media -- 10.5.1 Autoclavable Media -- 10.5.2 Sterile Filtration -- Protocol 10.10. Sterile Filtration with Syringe-Tip Filter -- Protocol 10.11. Sterile Filtration with Vacuum Filter Flask -- Protocol 10.12. Sterile Filtration with Small Inline Filter -- Protocol 10.13. Sterile Filtration with Large Inline Filter -- 10.5.3 Serum -- 10.5.4 Preparation and Sterilization of Other Reagents -- 10.6 Control, Testing, and Storage of Media -- 10.6.1 Quality Control -- 10.6.2 Sterility Testing -- 10.6.3 Culture Testing of Medium and Serum -- Protocol 10.14. Testing Medium by Plating Efficiency -- Protocol 10.15. Testing Medium by Growth -- 10.6.4 Storage -- References -- Suppliers -- 11. Primary Culture -- 11.1 Initiation of a Primary Cell Culture -- 11.1.1 Proteases Used in Disaggregation -- 11.1.2 Common Features of Disaggregation.
11.2 Isolation of the Tissue.
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Intro -- CULTURE OF ANIMAL CELLS -- Contents -- List of Figures -- List of Color Plates -- List of Tables -- List of Protocols -- List of Minireviews -- Preface and Acknlowledgments -- Abbreviations -- About the Companion Website -- 1. Introduction -- 1.1 Historical Background -- 1.2 Advantages of Tissue Culture -- 1.2.1 Control of the Environment -- 1.2.2 Characterization and Homogeneity of Sample -- 1.2.3 Economy, Scale, and Mechanization -- 1.2.4 In vitro Modeling of In vivo Conditions -- 1.3 Limitations -- 1.3.1 Expertise -- 1.3.2 Quantity -- 1.3.3 Dedifferentiation and Selection -- 1.3.4 Origin of Cells -- 1.3.5 Instability -- 1.4 Major Differences In Vitro -- 1.5 Types of Tissue Culture -- References -- 2. Biology of Cultured Cells -- 2.1 The Culture Environment -- 2.2 Cell Adhesion -- 2.2.1 Cell Adhesion Molecules -- 2.2.2 Intercellular Junctions -- 2.2.3 Cytoskeleton -- 2.2.4 Extracellular Matrix -- 2.2.5 Cell Motility -- 2.3 Cell Proliferation -- 2.3.1 Cell Cycle -- 2.3.2 Control of Cell Proliferation -- 2.4 Differentiation -- 2.4.1 Induction and Maintenance of Differentiation -- 2.4.2 Plasticity of Differentiation and Dedifferentiation -- 2.5 Cell Signaling -- 2.6 Energy Metabolism -- 2.7 Origin of Cultured Cells -- 2.7.1 Initiation of the Culture -- 2.7.2 Evolution of Cell Lines -- 2.7.3 Senescence -- 2.7.4 Transformation and the Development of Continuous Cell Lines -- 2.8 Definitions -- References -- Supplier -- 3. Laboratory Design and Layout -- 3.1 Planning, Furnishing, and Services -- 3.1.1 Requirements -- 3.1.2 Services -- 3.1.3 Ventilation -- 3.2 Design and Layout -- 3.2.1 Sterile Handling Area -- 3.2.2 Laminar Flow Hoods -- 3.2.3 Service Bench -- 3.2.4 Quarantine and Containment -- 3.2.5 Incubation -- 3.2.6 Preparation Area -- 3.2.7 Storage -- 3.3 Disaster Management -- References -- 4. Equipment and Materials.

4.1 Requirements of a Tissue Culture Laboratory -- 4.2 Sterile Working Area -- 4.2.1 Laminar-Flow BSC -- 4.2.2 Service Carts -- 4.2.3 Sterile Liquid Handling-Pipetting and Dispensing -- 4.2.4 Inverted Microscope -- 4.2.5 Camera and Monitor -- 4.2.6 Dissecting Microscope -- 4.2.7 Centrifuge -- 4.2.8 Cell Counting -- 4.3 Incubation and Culture -- 4.3.1 Incubator -- 4.3.2 Humid CO2 Incubator -- 4.3.3 Temperature Recorders -- 4.3.4 Roller Racks -- 4.3.5 Magnetic Stirrer -- 4.3.6 Culture Vessels -- 4.4 Preparation and Sterilization -- 4.4.1 Washup -- 4.4.2 Preparation of Media and Reagents -- 4.4.3 Sterilization -- 4.5 Storage -- 4.5.1 Consumables -- 4.5.2 Refrigerators and Freezers -- 4.5.3 Cryostorage Containers -- 4.5.4 Controlled-Rate Freezer -- 4.6 Supplementary Laboratory Equipment -- 4.6.1 Computers and Networks -- 4.6.2 Upright Microscope -- 4.6.3 Low-Temperature Freezer -- 4.6.4 Confocal Microscope -- 4.6.5 PCR Thermal Cycler -- 4.7 Specialized Equipment -- 4.7.1 Microinjection Facilities -- 4.7.2 Colony Counter -- 4.7.3 Centrifugal Elutriator -- 4.7.4 Flow Cytometer -- References -- Suppliers -- 5. Aseptic Technique -- 5.1 Objectives of Aseptic Technique -- 5.1.1 Risk of Contamination -- 5.1.2 Maintaining Sterility -- 5.2 Elements of Aseptic Environment -- 5.2.1 Laminar Flow -- 5.2.2 Quiet Area -- 5.2.3 Work Surface -- 5.2.4 Personal Hygiene -- 5.2.5 Reagents and Media -- 5.2.6 Cultures -- 5.3 Sterile Handling -- 5.3.1 Swabbing -- 5.3.2 Capping -- 5.3.3 Flaming -- 5.3.4 Handling Bottles and Flasks -- 5.3.5 Pipetting -- 5.3.6 Large-Volume Dispensing -- 5.3.7 Pouring -- 5.4 Standard Procedure -- Protocol 5.1. Aseptic Technique In BSC -- Protocol 5.2. Working On The Open Bench -- 5.4.1 Culture Flasks and Bottles -- 5.4.2 Petri Dishes and Multiwell Plates -- Protocol 5.3. Handling Dishes or Plates -- 5.5 Apparatus and Equipment.

5.5.1 Refrigerators and Coldrooms -- 5.5.2 Incubators -- Protocol 5.4. Cleaning Incubators -- 5.5.3 Boxed Cultures -- 5.5.4 Gassing with CO2 -- References -- Suppliers -- 6. Safety, Bioethics, and Validation -- 6.1 Laboratory Safety -- 6.2 Risk Assessment -- 6.3 Standard Operating Procedures -- 6.4 Safety Regulations -- 6.5 General Safety -- 6.5.1 Operator -- 6.5.2 Equipment -- 6.5.3 Glassware and Sharp Items -- 6.5.4 Chemical Toxicity -- 6.5.5 Gases -- 6.5.6 Liquid Nitrogen -- 6.5.7 Burns -- 6.6 Fire -- 6.7 Ionizing Radiation -- 6.7.1 Ingestion -- 6.7.2 Disposal of Radioactive Waste -- 6.7.3 Irradiation from Labeled Reagents -- 6.7.4 Irradiation from High-Energy Sources -- 6.8 Biohazards -- 6.8.1 Levels of Biological Containment -- 6.8.2 Biological Safety Cabinets (BSCs) -- 6.8.3 Human Biopsy Material -- 6.8.4 Cell Lines -- 6.8.6 Genetic Manipulation -- 6.8.6 Disposal of Biohazardous Waste -- 6.8.7 Decontamination and Fumigation -- 6.9 Bioethics -- 6.9.1 Animal Tissue -- 6.9.2 Human Tissue -- 6.10 Quality Assurance -- 6.10.1 Procedures -- 6.10.2 Quality Control (QC) -- 6.10.3 Validation -- 6.10.4 Authentication -- 6.10.5 Provenance -- 6.10.6 Contamination -- References -- Suppliers -- 7. Culture Vessels and Substrates -- 7.1 The Substrate -- 7.1.1 Attachment and Growth -- 7.1.2 Common Substrate Materials -- 7.1.3 Alternative Substrates -- 7.2 Treated Surfaces -- 7.2.1 Substrate Coating -- Protocol 7.1. Preparation of ECM -- 7.2.2 Feeder Layers -- 7.2.3 Nonadhesive Substrates -- 7.3 Choice of Culture Vessel -- 7.3.1 Cell Yield -- 7.3.2 Multiwell Plates -- 7.3.3 Flasks and Petri Dishes -- 7.3.4 High Yields -- 7.3.5 Suspension Culture -- 7.3.6 Venting -- 7.3.6 Sampling and Analysis -- 7.3.7 Uneven Growth -- 7.3.8 Cost -- 7.4 Specialized Systems -- 7.4.1 Permeable Supports -- 7.4.2 Three-Dimensional Matrices -- References -- Suppliers.

8. Defined Media and Supplements -- 8.1 D evelopment of Media -- 8.2 Physicochemical Properties -- 8.2.1 pH -- Protocol 8.1. Preparation of pH Standards -- 8.2.2 CO2 and Bicarbonate -- 8.2.3 Buffering -- 8.2.4 Oxygen -- Minireview 8.1. Hypoxic Cell Culture -- 8.2.5 Osmolality -- 8.2.6 Temperature -- 8.2.7 Viscosity -- 8.2.8 Surface Tension and Foaming -- 8.3 Balanced Salt Solutions -- 8.4 Complete Media -- 8.4.1 Amino Acids -- 8.4.2 Vitamins -- 8.4.3 Salts -- 8.4.4 Glucose -- 8.4.5 Organic Supplements -- 8.4.6 Hormones and Growth Factors -- 8.4.7 Antibiotics -- 8.5 Serum -- 8.5.1 Protein -- 8.5.2 Growth Factors -- 8.5.3 Hormones -- 8.5.4 Nutrients and Metabolites -- 8.5.5 Lipids -- 8.5.6 Minerals -- 8.5.7 Inhibitors -- 8.6 Selection of Medium and Serum -- 8.6.1 Serum Batch Reservation -- 8.6.2 Testing Serum -- 8.6.3 Heat Inactivation -- 8.7 Other Supplements -- 8.7.1 Amino Acid Hydrolysates -- 8.7.2 Embryo Extract -- 8.7.3 Conditioned Medium -- 8.8 Storage -- References -- Suppliers -- 9. Serum-Free Media -- 9.1 Disadvantages of Serum -- 9.2 Advantages of Serum-Free Media -- 9.3 Disadvantages of Serum-Free Media -- 9.4 Replacement of Serum -- 9.4.1 Commercially Available Serum-Free Media -- 9.4.2 Serum Replacements -- 9.4.3 Serum-Free Subculture -- 9.4.4 Hormones -- 9.4.5 Growth Factors -- 9.4.6 Nutrients in Serum -- 9.4.7 Proteins and Polyamines -- 9.4.8 Viscosity -- 9.5 D evelopment of Serum-Free Medium -- 9.6 Selection of Serum-Free Medium -- 9.6.1 Cell or Product Specificity -- 9.6.2 Adaptation of Cell Lines to Serum-Free Media -- 9.7 Preparation of Serum-Free Medium -- 9.8 Animal Protein-Free Media -- 9.9 Conclusions -- References -- Suppliers -- 10. Preparation and Sterilization -- 10.1 Preparation of Reagents and Materials -- 10.2 Sterilization of Apparatus and Liquids -- 10.2.1 Dry-Heat Sterilization -- 10.2.2 Pressurized-Steam Sterilization.

10.2.3 Irradiation -- 10.2.4 Chemical Sterilization -- 10.2.5 Sterility Indicators -- 10.2.6 Filter Sterilization -- 10.3 Apparatus -- 10.3.1 Glassware -- Protocol 10.1. Preparation and Sterilization of Glassware -- 10.3.2 Glass Pipettes -- Protocol 10.2. Preparation and Sterilization of Glass Pipettes -- 10.3.3 Screw Caps -- Protocol 10.3. Preparation and Sterilization of Screw Caps -- 10.3.4 Selection of Detergent -- 10.3.5 Miscellaneous Equipment -- 10.3.6 Reusable Sterilizing Filters -- Protocol 10.4. Sterilizing Filter Assemblies -- 10.4 Reagents and Media -- 10.4.1 Water -- Protocol 10.5. Preparation and Sterilization of Ultrapure Water (UPW) -- 10.4.2 Maintenance of Water Purifier -- 10.4.3 Balanced Salt Solutions -- Protocol 10.6. Preparation and Sterilization of D-PBSA -- 10.4.4 Preparation and Sterilization of Media -- Protocol 10.7. Preparation of Medium from 1× Stock -- Protocol 10.8. Preparation of Medium from 10× Concentrate -- Protocol 10.9. Preparation of Medium from Powder -- 10.5 Sterilization of Media -- 10.5.1 Autoclavable Media -- 10.5.2 Sterile Filtration -- Protocol 10.10. Sterile Filtration with Syringe-Tip Filter -- Protocol 10.11. Sterile Filtration with Vacuum Filter Flask -- Protocol 10.12. Sterile Filtration with Small Inline Filter -- Protocol 10.13. Sterile Filtration with Large Inline Filter -- 10.5.3 Serum -- 10.5.4 Preparation and Sterilization of Other Reagents -- 10.6 Control, Testing, and Storage of Media -- 10.6.1 Quality Control -- 10.6.2 Sterility Testing -- 10.6.3 Culture Testing of Medium and Serum -- Protocol 10.14. Testing Medium by Plating Efficiency -- Protocol 10.15. Testing Medium by Growth -- 10.6.4 Storage -- References -- Suppliers -- 11. Primary Culture -- 11.1 Initiation of a Primary Cell Culture -- 11.1.1 Proteases Used in Disaggregation -- 11.1.2 Common Features of Disaggregation.

11.2 Isolation of the Tissue.

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