Details

Freshney's Culture of Animal Cells


Freshney's Culture of Animal Cells

A Manual of Basic Technique and Specialized Applications
8. Aufl.

von: Amanda Capes-Davis, R. Ian Freshney

86,99 €

Verlag: Wiley-Blackwell
Format: EPUB
Veröffentl.: 17.03.2021
ISBN/EAN: 9781119513049
Sprache: englisch
Anzahl Seiten: 832

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Beschreibungen

<b>FRESHNEY’S CULTURE OF ANIMAL CELLS</b> <p><b>THE NEW EDITION OF THE LEADING TEXT ON THE BASIC METHODOLOGY OF CELL CULTURE, FULLY UPDATED TO REFLECT NEW APPLICATIONS INCLUDING IPSCS, CRISPR, AND ORGAN-ON-CHIP TECHNOLOGIES</b> <p><i>Freshney’s Culture of Animal Cells</i> is the most comprehensive and up-to-date resource on the principles, techniques, equipment, and applications in the field of cell and tissue culture. Explaining both how to do tissue culture and why a technique is done in a particular way, this classic text covers the biology of cultured cells, how to select media and substrates, regulatory requirements, laboratory protocols, aseptic technique, experimental manipulation of animal cells, and much more. <p>The eighth edition contains extensively revised material that reflects the latest techniques and emerging applications in cell culture, such as the use of CRISPR/Cas9 for gene editing and the adoption of chemically defined conditions for stem cell culture. A brand-new chapter examines the origin and evolution of cell lines, joined by a dedicated chapter on irreproducible research, its causes, and the importance of reproducibility and good cell culture practice. Throughout the book, updated chapters and protocols cover topics including live-cell imaging, 3D culture, scale-up and automation, microfluidics, high-throughput screening, and toxicity testing. This landmark text: <li><bl>Provides comprehensive single-volume coverage of basic skills and protocols, specialized techniques and applications, and new and emerging developments in the field</bl></li> <li><bl>Covers every essential area of animal cell culture, including lab design, disaster and contingency planning, safety, bioethics, media preparation, primary culture, mycoplasma and authentication testing, cell line characterization and cryopreservation, training, and troubleshooting</bl></li> <li><bl>Features a wealth of new content including protocols for gene delivery, iPSC generation and culture, and tumor spheroid formation</bl></li> <li><bl>Includes an updated and expanded companion website containing figures, artwork, and supplementary protocols to download and print</bl></li> <p>The eighth edition of <i>Freshney’s Culture of Animal Cells</i> is an indispensable volume for anyone involved in the field, including undergraduate and graduate students, clinical and biopharmaceutical researchers, bioengineers, academic research scientists, and managers, technicians, and trainees working in cell biology, molecular biology, and genetics laboratories.
<p>Foreword xix</p> <p>Acknowledgments xxi</p> <p>Abbreviations xxiii</p> <p>Book Navigation xxix</p> <p><b>Part I Understanding Cell Culture 1</b></p> <p><b>1. Introduction 3</b></p> <p>1.1 Terminology 3</p> <p>1.2 Historical Development 4</p> <p>1.3 Applications 12</p> <p>1.4 Advantages of Tissue Culture 13</p> <p>1.5 Limitations of Tissue Culture 15</p> <p>References 18</p> <p><b>2. Biology of Cultured Cells 23</b></p> <p>2.1 The Culture Environment 23</p> <p>2.2 Cell Adhesion 23</p> <p>2.3 Cell Division 28</p> <p>2.4 Cell Fate 30</p> <p>2.5 Cell Death 35</p> <p>References 36</p> <p><b>3. Origin and Evolution of Cultured Cells 39</b></p> <p>3.1 Origin of Cultured Cells 39</p> <p>3.2 Evolution of Cell Lines 40</p> <p>3.3 Changes in Genotype 43</p> <p>3.4 Changes in Phenotype 46</p> <p>3.5 Senescence and Immortalization 48</p> <p><i>Minireview M3.1 Senescence and Immortalization 48</i></p> <p>3.6 Transformation 50</p> <p>3.7 Conclusions: Origin and Evolution 58</p> <p>References 58</p> <p><b>Part II Laboratory and Regulatory Requirements 63</b></p> <p><b>4. Laboratory Design and Layout 65</b></p> <p>4.1 Design Requirements 65</p> <p>4.2 Layout of Laboratory Areas 74</p> <p>4.3 Disaster and Contingency Planning 80</p> <p>References 83</p> <p><b>5. Equipment and Materials 85</b></p> <p>5.1 Sterile Handling Area Equipment 85</p> <p>5.2 Imaging and Analysis Equipment 97</p> <p>5.3 Incubation Equipment 99</p> <p>5.4 Preparation and Washup Equipment 104</p> <p>5.5 Cold Storage Equipment 107</p> <p>References 109</p> <p><b>6. Safety and Bioethics 111</b></p> <p>6.1 Laboratory Safety 111</p> <p>6.2 Hazards in Tissue Culture Laboratories 117</p> <p>6.3 Biosafety 121</p> <p>6.4 Bioethics 129</p> <p>References 132</p> <p><b>7. Reproducibility and Good Cell Culture Practice 137</b></p> <p>7.1 Reproducibility 137</p> <p>7.2 Good Practice Requirements 141</p> <p>7.3 Cell Line Provenance 145</p> <p>7.4 Validation Testing 146</p> <p>7.5 Quality Assurance (QA) 148</p> <p>7.6 Replicate Sampling 150</p> <p>References 151</p> <p><b>Part III Medium and Substrate Requirements 155</b></p> <p><b>8. Culture Vessels and Substrates 157</b></p> <p>8.1 Attachment and Growth Requirements 157</p> <p>8.2 Substrate Materials 158</p> <p>8.3 Substrate Treatments 159</p> <p>8.4 Feeder Layers 163</p> <p>8.5 Choice of Culture Vessel 164</p> <p>8.6 Application-Specific Vessels 170</p> <p>References 173</p> <p><b>9. Defined Media and Supplements 177</b></p> <p>9.1 Medium Development 177</p> <p>9.2 Physicochemical Properties 177</p> <p>9.3 Balanced Salt Solutions 185</p> <p>9.4 Media Formulations 186</p> <p>9.5 Serum 189</p> <p>9.6 Other Media Supplements 191</p> <p>9.7 Choice of Complete Medium 191</p> <p>9.8 Storage of Medium and Serum 194</p> <p>Suppliers 194</p> <p>References 194</p> <p><b>10. Serum-Free Media 199</b></p> <p>10.1 Rationale for Serum-Free Medium 199</p> <p>10.2 Development of Serum-Free Medium 201</p> <p>10.3 Serum-Free Media Formulations 202</p> <p>10.4 Serum-Free Supplements 203</p> <p>10.5 Serum Replacements 209</p> <p>10.6 Use of Serum-Free Medium 209</p> <p>10.7 Xeno-Free Media 213</p> <p>10.8 Animal Product-Free Media 214</p> <p>10.9 Conclusions: Serum-Free Media 214</p> <p>Suppliers 214</p> <p>References 215</p> <p><b>11. Preparation and Sterilization 219</b></p> <p>11.1 Terminology: Preparation 219</p> <p>11.2 Sterilization Methods 220</p> <p>11.3 Glassware 224</p> <p><i>Protocol P11.1 Preparation and Sterilization of Glassware 224</i></p> <p>11.4 Other Laboratory Apparatus 229</p> <p>11.5 Water 229</p> <p>11.6 Media and Other Reagents 233</p> <p>11.7 Sterile Filtration 238</p> <p>11.8 Medium Testing 242</p> <p>Suppliers 247</p> <p>References 247</p> <p><b>Part IV Handling Cultures 249</b></p> <p><b>12. Aseptic Technique 251</b></p> <p>12.1 Objectives of Aseptic Technique 251</p> <p>12.2 Elements of Aseptic Environment 252</p> <p>12.3 Sterile Handling 258</p> <p>12.4 Good Aseptic Technique 260</p> <p>12.5 Controlling Equipment Contamination 265</p> <p>Suppliers 267</p> <p>References 267</p> <p><b>13. Primary Culture 269</b></p> <p>13.1 Rationale for Primary Culture 269</p> <p>13.2 Initiation of Primary Culture 270</p> <p>13.3 Tissue Acquisition and Isolation 274</p> <p>13.4 Primary Explantation 281</p> <p><i>Protocol P13.3 Culture of Primary Explants 281</i></p> <p>13.5 Enzymatic Disaggregation 283</p> <p>13.6 Mechanical Disaggregation 290</p> <p><i>Protocol P13.7 Mechanical Disaggregation by Sieving 291</i></p> <p>13.7 Enrichment of Viable Cells 292</p> <p><i>Protocol P13.8 Enrichment of Viable Cells 292</i></p> <p>13.8 Record Keeping for Primary Culture 293</p> <p>13.9 Conclusions: Primary Culture 294</p> <p>Suppliers 294</p> <p>References 294</p> <p><b>14. Subculture and Cell Lines 297</b></p> <p>14.1 Terminology: Cell Line and Subculture 297</p> <p>14.2 Initiating a Cell Line 298</p> <p>14.3 Choosing a Cell Line 300</p> <p>14.4 Maintaining a Cell Line 304</p> <p>14.5 Replacing Medium (Feeding) 309</p> <p>14.6 Subculture (Passaging) 312</p> <p>14.7 Maintaining Suspension Cultures 320</p> <p>14.8 Serum-Free Subculture 322</p> <p>14.9 Record Keeping for Cell Lines 323</p> <p>Suppliers 324</p> <p>References 325</p> <p><b>15. Cryopreservation and Banking 327</b></p> <p>15.1 Principles of Cryopreservation 327</p> <p>15.2 Apparatus for Cryopreservation 329</p> <p>15.3 Requirements for Cryopreservation 335</p> <p>15.4 Cryopreservation Procedures 336</p> <p>15.5 Cell Banking Procedures 341</p> <p>15.6 Cell Repositories 342</p> <p>15.7 Record Keeping for Frozen Stocks 345</p> <p>15.8 Transporting Cells 347</p> <p>Suppliers 348</p> <p>References 348</p> <p><b>Part V Validation and Characterization 351</b></p> <p><b>16. Microbial Contamination 353</b></p> <p>16.1 Sources of Contamination 353</p> <p>16.2 Management of Contamination 359</p> <p><i>Protocol P16.1 Disposal of Contaminated Cultures 360</i></p> <p>16.3 Visible Microbial Contamination 361</p> <p>16.4 Mycoplasma Contamination 364</p> <p>16.5 Viral Contamination 373</p> <p>16.6 Dealing with Persistent Contamination 376</p> <p>Suppliers 376</p> <p>References 376</p> <p><b>17. Cell Line Misidentification and Authentication 381</b></p> <p>17.1 Terminology: Cross-Contamination, Misidentification, and Authentication 381</p> <p>17.2 Misidentified Cell Lines 382</p> <p>17.3 Cell Line Authentication 386</p> <p>17.4 Authentication of Challenging Samples 401</p> <p>17.5 Conclusions: Authentication 403</p> <p>Suppliers 403</p> <p>References 403</p> <p><b>18. Cell Line Characterization 409</b></p> <p>18.1 Priorities and Essential Characterization 409</p> <p>18.2 Genotype-Based Characterization 416</p> <p>18.3 Phenotype-Based Characterization 419</p> <p>18.4 Cell Imaging 423</p> <p>18.5 Cell Staining 428</p> <p>Suppliers 430</p> <p>References 430</p> <p><b>19. Quantitation and Growth Kinetics 437</b></p> <p>19.1 Cell Counting 437</p> <p>19.3 Cell Proliferation 450</p> <p>19.4 Cloning Efficiency 456</p> <p>19.5 DNA Synthesis 460</p> <p>19.6 Cell Cycle Analysis 461</p> <p>Suppliers 461</p> <p>References 461</p> <p><b>Part VI Physical and Genetic Manipulation 465</b></p> <p><b>20. Cell Cloning and Selection 467</b></p> <p>20.1 Terminology: Cloning and Selection 467</p> <p>20.2 Cloning by Limiting Dilution 468</p> <p>20.3 Cloning in Suspension 473</p> <p>20.4 Selection of Clones 477</p> <p>20.5 Replica Plating 480</p> <p>20.6 Stimulation of Cloning Efficiency 481</p> <p>20.7 Selective Culture Conditions 485</p> <p>20.8 Conclusions: Cloning and Selection 487</p> <p>Suppliers 487</p> <p>References 487</p> <p><b>21. Cell Separation and Sorting 491</b></p> <p>21.1 Cell Density and Isopycnic Centrifugation 491</p> <p>21.2 Cell Size and Sedimentation Velocity 495</p> <p>21.3 Magnetic Separation and Sorting 496</p> <p><i>Protocol P21.2 Magnet-Activated Cell Sorting (MACS) 499</i></p> <p>21.4 Fluorescence-Activated Cell Sorting (FACS) 500</p> <p>21.5 Microfluidic Sorting 502</p> <p><i>Minireview M21.1 Microfluidic Cell Culture 503</i></p> <p>21.6 Conclusions: Sorting and Separation 505</p> <p>Suppliers 505</p> <p>References 505</p> <p><b>22. Genetic Modification and Immortalization 509</b></p> <p>22.1 Gene Delivery 509</p> <p>22.2 Gene Editing 517</p> <p>22.3 Immortalization 523</p> <p>22.4 Screening and Artifacts 526</p> <p>Suppliers 528</p> <p>References 528</p> <p><b>Part VII Stem Cells and Differentiated Cells 535</b></p> <p><b>23. Culture of Stem Cells 537</b></p> <p>23.1 Terminology: Stem Cells 537</p> <p>23.2 Embryonic Stem Cells (ESCs) 540</p> <p>23.3 Induction of Pluripotency 545</p> <p><i>Protocol P23.1 Generation of iPSCs Using Sendai Viral Vectors 547</i></p> <p>23.4 Human Pluripotent Stem Cell (hPSC) Lines 549</p> <p>23.5 Perinatal Stem Cells 556</p> <p>23.6 Adult Stem Cells 557</p> <p>23.7 Stem Cell Characterization and Banking 558</p> <p>23.8 Conclusions: Culture of Stem Cells 560</p> <p>Suppliers 561</p> <p>References 561</p> <p><b>24. Culture of Specific Cell Types 567</b></p> <p>24.1 Specialized Cells and Their Availability 567</p> <p>24.2 Epithelial Cells 572</p> <p>24.3 Mesenchymal Cells 577</p> <p>24.4 Neuroectodermal Cells 580</p> <p>24.5 Hematopoietic Cells 581</p> <p>24.6 Culture of Cells from Poikilotherms 585</p> <p>Suppliers 587</p> <p>References 587</p> <p><b>25. Culture of Tumor Cells 593</b></p> <p>25.1 Challenges of Tumor Cell Culture 593</p> <p>25.2 Primary Culture of Tumor Cells 594</p> <p>25.3 Development of Tumor Cell Lines 596</p> <p>25.4 Selective Culture of Tumor Cells 599</p> <p>25.5 Specific Tumor Types 603</p> <p>25.6 Cancer Stem Cells (CSCs) 606</p> <p><i>Minireview M25.1 Culture of Cancer Stem Cells 606</i></p> <p>Suppliers 608</p> <p>References 608</p> <p><b>26. Differentiation 615</b></p> <p>26.1 <i>In Vitro </i>Models of Differentiation 615</p> <p>26.2 Differentiation Status in Culture 617</p> <p>26.3 Induction of Differentiation 620</p> <p>26.4 Practical Aspects 628</p> <p>26.5 Ongoing Challenges 629</p> <p>Suppliers 631</p> <p>References 631</p> <p><b>Part VIII Model Environments and Applications 639</b></p> <p><b>27. Three-Dimensional Culture 641</b></p> <p>27.1 Terminology: 3D Culture 641</p> <p>27.2 Technologies for 3D Culture 643</p> <p><i>Minireview M27.1 Advances in Technologies Enabling 3D Cell Culture and the Formation of Tissue-Like Architecture </i>In Vitro<i> 643</i></p> <p>27.3 Benefits and Limitations of 3D Culture 646</p> <p>27.4 Scaffold-Free 3D Culture Systems 647</p> <p>27.5 Scaffold-Based 3D Culture Systems 652</p> <p>27.6 Organoid Culture 659</p> <p>27.7 Organotypic Culture 660</p> <p>27.8 Organ Culture 662</p> <p>27.9 Characterization of 3D Cultures 662</p> <p>Suppliers 663</p> <p>References 663</p> <p><b>28. Scale-Up and Automation 669</b></p> <p>28.1 Terminology: Scale-Up and Bioreactors 669</p> <p>28.2 Scale-Up in Suspension 671</p> <p>28.3 Scale-Up in Monolayer 677</p> <p>28.4 Monitoring and Process Control 685</p> <p>28.5 Scale-Up for Manufacture 688</p> <p><i>Minireview M28.1 Culture Scale-Up and Bioreactors 688</i></p> <p>28.6 High-Throughput Screening 691</p> <p>28.7 Automation and Bioprinting 691</p> <p>Suppliers 696</p> <p>References 696</p> <p><b>29. Toxicity Testing 701</b></p> <p>29.1 <i>In Vitro </i>Toxicity Testing 701</p> <p>29.2 Cytotoxicity Assays 704</p> <p>29.3 Genotoxicity Assays 715</p> <p>29.4 Carcinogenicity Assays 716</p> <p>29.5 Advanced Models for Toxicity Testing 716</p> <p>Suppliers 719</p> <p>References 719</p> <p><b>Part IX Teaching and Troubleshooting 725</b></p> <p><b>30. Training 727</b></p> <p>30.1 Training Principles 727</p> <p>30.2 Training Programs 729</p> <p>References 731</p> <p><b>31. Problem Solving 733</b></p> <p>31.1 Microbial Contamination 733</p> <p>31.2 Cross-Contamination and Misidentification 737</p> <p>31.3 Chemical Contamination 738</p> <p>31.4 Slow Cell Growth 738</p> <p>31.5 Abnormal Cell Appearance 740</p> <p>31.6 Problems with Materials 741</p> <p>31.7 Problems with Primary Culture 744</p> <p>31.8 Problems with Feeding or Subculture 746</p> <p>31.9 Problems with Cryopreservation 748</p> <p>31.10 Problems with Cloning 750</p> <p>References 752</p> <p><b>32. In Conclusion 753</b></p> <p><b>Appendix A Glossary 755</b></p> <p><b>Appendix B Calculations and Preparation of Reagents 761</b></p> <p>Calculations 761</p> <p>Counting Cells with a Hemocytometer 761</p> <p>Dilution of a Cell Suspension 761</p> <p>Population Doubling Level (PDL) 761</p> <p>Molarity 762</p> <p>Percentages and Dilutions 762</p> <p>Pressure 762</p> <p>Rotor Speed (rpm to g) 762</p> <p>Preparation of Reagents 762</p> <p>Acetic Acid: Methanol 762</p> <p>Agar (2.5%) 762</p> <p>Alcohol (70%) 762</p> <p>Bacto<sup>™</sup> Peptone (5%) 763</p> <p>Balanced Salt Solutions 763</p> <p>Carboxymethylcellulose (CMC; 4%) 763</p> <p>Chick Embryo Extract 763</p> <p>Collagenase 763</p> <p>Collection Medium 763</p> <p>Crystal Violet (0.1%) 764</p> <p>Dexamethasone (1 mg/ml) 764</p> <p>Dissection Balanced Salt Solution (DBSS) 764</p> <p>Dulbecco’s Phosphate-Buffered Saline Without Ca<sup>2</sup><sup>+</sup> and Mg<sup>2</sup><sup>+</sup> (DPBS-A) 764</p> <p>EDTA (10 mM in DPBS-A) 764</p> <p>EGTA 764</p> <p>Erythrosin B 764</p> <p>Gelatin (1%) 765</p> <p>Giemsa Stain 765</p> <p>Glucose (20%) 765</p> <p>Glutamine 200 mM 765</p> <p>Hanks’s Balanced Salt Solution (HBSS) 765</p> <p>HAT Medium 765</p> <p>HB Medium 765</p> <p>HEPES 765</p> <p>Hoechst 33258 766</p> <p>Media 766</p> <p>2-Mercaptoethanol (𝛽-Mercaptoethanol; 0.1 M) 766</p> <p>Methylcellulose (Methocel, 1.6%) 766</p> <p>Mitomycin C (100 μg/ml) 766</p> <p>MTT (50 mg/ml) 766</p> <p>N2 Supplement 766</p> <p>N2B27 Medium 767</p> <p>Naphthalene Black (Amido Black; 1%) 767</p> <p>Non-essential Amino Acids (NEAA, 100×) 767</p> <p>Paraformaldehyde (4%) 767</p> <p>Trypan Blue (0.4%) 767</p> <p>Trypsin (2.5%) 768</p> <p>Versene 768</p> <p>Suppliers 768</p> <p>References 768</p> <p><b>Appendix C Media Formulations 769</b></p> <p>References 779</p> <p>Index 781 </p>
<p><b>AMANDA CAPES-DAVIS, PHD,</b> is a cell culture scientist and technical writer. She was Founding Manager and Honorary Scientist at CellBank Australia, Children’s Medical Research Institute (CMRI), and is a member of the International Cell Line Authentication Committee (ICLAC). She was a Reviewing Editor for the 7th edition of <i>Culture of Animal Cells</i>, and has written numerous journal articles, policies, protocols, and white papers on good cell culture practice.</p> <p><b>R. IAN FRESHNEY, PHD,</b> was an honorary Senior Research Fellow at the Institute of Cancer Sciences at the University of Glasgow, UK. Dr Freshney, who died in 2019, was a world-renowned cancer biologist and a pioneer in cell culture techniques who made important contributions to new approaches for treating cancer patients. He taught cell culture courses at national and international level, and wrote and edited numerous books, including the first seven editions of <i>Culture of Animal Cells</i>.
<p><b>THE NEW EDITION OF THE LEADING TEXT ON THE BASIC METHODOLOGY OF CELL CULTURE, FULLY UPDATED TO REFLECT NEW APPLICATIONS INCLUDING IPSCS, CRISPR, AND ORGAN-ON-CHIP TECHNOLOGIES</b> <p><i>Freshney’s Culture of Animal Cells</i> is the most comprehensive and up-to-date resource on the principles, techniques, equipment, and applications in the field of cell and tissue culture. Explaining both how to do tissue culture and why a technique is done in a particular way, this classic text covers the biology of cultured cells, how to select media and substrates, regulatory requirements, laboratory protocols, aseptic technique, experimental manipulation of animal cells, and much more. <p>The eighth edition contains extensively revised material that reflects the latest techniques and emerging applications in cell culture, such as the use of CRISPR/Cas9 for gene editing and the adoption of chemically defined conditions for stem cell culture. A brand-new chapter examines the origin and evolution of cell lines, joined by a dedicated chapter on irreproducible research, its causes, and the importance of reproducibility and good cell culture practice. Throughout the book, updated chapters and protocols cover topics including live-cell imaging, 3D culture, scale-up and automation, microfluidics, high-throughput screening, and toxicity testing. This landmark text: <li><bl>Provides comprehensive single-volume coverage of basic skills and protocols, specialized techniques and applications, and new and emerging developments in the field</bl></li> <li><bl>Covers every essential area of animal cell culture, including lab design, disaster and contingency planning, safety, bioethics, media preparation, primary culture, mycoplasma and authentication testing, cell line characterization and cryopreservation, training, and troubleshooting</bl></li> <li><bl>Features a wealth of new content including protocols for gene delivery, iPSC generation and culture, and tumor spheroid formation</bl></li> <li><bl>Includes an updated and expanded companion website containing figures, artwork, and supplementary protocols to download and print</bl></li> <p>The eighth edition of <i>Freshney’s Culture of Animal Cells</i> is an indispensable volume for anyone involved in the field, including undergraduate and graduate students, clinical and biopharmaceutical researchers, bioengineers, academic research scientists, and managers, technicians, and trainees working in cell biology, molecular biology, and genetics laboratories.

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