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Basic Cell Culture

ISBN-10: 0199638535

ISBN-13: 9780199638536

Edition: 2nd 2001 (Revised)

Authors: John Davis

List price: $125.00
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Description:

The use of animal, including human, cell culture has expanded enormously during the last 25 years, with new applications appearing all the time. The aim of this book is to guide the newcomer progressively through all those areas which nowadays are basic to the performance of cell culture. The book should also prove useful to the experienced worker when entering a new field, setting up a new laboratory, or as a source of reference on basic techniques. Revised and updated chapters cover topics such as equipping a cell culture laboratory, sterilization, culture media, basic culture technique, cell cloning, the isolation of new cell lines, contamination procedures, and good laboratory practice. There is a brand new chapter on the microscopy of living cells, and an appendix of online resources for cell biology.
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Book details

List price: $125.00
Edition: 2nd
Copyright year: 2001
Publisher: Oxford University Press, Incorporated
Publication date: 3/21/2002
Binding: Paperback
Pages: 408
Size: 7.50" wide x 9.75" long x 0.75" tall
Weight: 1.980
Language: English

List of protocols
Abbreviations
The cell culture laboratory
Animal cell culture: past, present, and future
The laboratory
Design concepts and layout
Services
Equipping the laboratory
Major items of equipment and instrumentation
Culture plasticware and associated small consumable items
Washing reusable tissue culture equipment
Soaking
Washing
Pipette washing
General care and maintenance of the tissue culture laboratory
Acknowledgements
References
Sterilization
Introduction
What is sterilization?
The importance of sterility in cell culture
The use of antibiotics
Basic principles
Wet heat at up to 100[degree]C
Theory
Pasteurization
100[degree]C
Wet heat above 100[degree]C and autoclaving
Theory
Steam
Types of autoclave
Preparation of the load and operation of the autoclave
Autoclave testing
Dry heat
Incineration
Hot-air ovens
Irradiation
Ultraviolet light
Gamma rays
Chemical sterilization
Fumigation
Liquid disinfectants
Filtration
Filters for bacteria and fungi
Filters for viruses
HEPA filters
Viruses and prions
Virus elimination
Prions
Safety in the laboratory
Good Manufacturing Practice and Good Laboratory Practice
Acknowledgements
References
Culture media
Introduction
Basal media
Types of basal media
Constituents of basal media
Choice of basal medium
Preparation of basal medium
Serum
Why use serum?
Types of serum
Constituents of serum
Potential problems with the use of serum
Sourcing and selection of serum
Serum storage and use
Replacement of serum in media
Reduced-serum media for early-passage normal human cells
Serum-free media (SFM)
Design of SFM
Influence of cell culture systems on the choice of medium
Batch or perfusion cultures
Anchorage-dependent cells
Stirred suspension cultures
High density culture systems
Very low density cultures
In-house medium development and production versus commercial supply
Economic considerations
Development of a dedicated medium
Quality assurance and control
References
Microscopy of living cells
Introduction
The modern light microscope
Basic scheme
Properties of light
Magnification
Resolution
Aberrations and their corrections
Illumination
Maintenance
Microscopic techniques
Bright field
Oblique illumination
Dark field
Phase-contrast
Differential interference contrast (DIC)
Hoffman modulation contrast
Reflection contrast
Varel contrast
Polarizing microscopy
Interference microscopy
Fluorescence microscopy
Specimen preparation
Culture flasks and Petri dishes
Coverslips
Chambers
Interaction with microscopic techniques
Environmental control for microscopic specimens
Recording of microscopic images
Time-lapse imaging
Analysis of microscopic images
Image enhancement by deconvolution
Image processing--extraction of morphometric data
Data analysis--statistical evaluation
Data interpretation--simulation modelling
Acknowledgements
References
Basic cell culture technique and the maintenance of cell lines
Introduction
The terminology of cell and tissue culture
Basic components of the cell culture environment
Culture medium
Physicochemical factors
Stationary versus dynamic media supply
Sterile technique and contamination control
Working within the laminar flow hood
Pipetting and prevention of aerosol formation
Additional considerations for good sterile technique
Prevention of cellular cross-contamination
Cell culture at the open bench
General procedures for the cell culture laboratory
Maintenance of the laboratory
Daily inspection of ongoing cultures
Maintenance of stock cultures
The culture of attached cells
Routine culture substrates
Choice of culture vessels
Artefacts of cell attachment and growth
Use of attachment factors and bio-substrates
Alternative culture substrates
Three-dimensional matrices
Culture of cells on microcarriers
Mass culture systems for adherent cells
The culture of cells in suspension
Non-adherent substrates for small-scale culture
Mass culture of cells in fluid suspension
Micro-encapsulation
In vitro cell growth behaviour
Adaptation to culture
Phases of cell growth
Determinations of cell growth data
Calculation of in vitro age
Multiplication rate and population doubling time
Counting cells in suspension
Counting cells adherent to a substrate
Phases of the cell cycle
Cell generation time by time-lapse photomicrography
Growth fraction
Expressions of culture 'efficiency'
Cryopreservation and retrieval of cells from frozen storage
Purpose of cell banking
Mechanism of cell freezing, and factors that affect viability
Supplies and equipment for cell freezing
Additional comments regarding cryopreservation
Transportation of cells
Transporting frozen cells
Transporting growing cells
Safety in the cell culture laboratory
Potential risks in routine cell culture
Awareness of increased risk associated with human cells
Classification of cell lines as aetiological agents
Precautions in handling pathogenic organisms and human cells
References
Primary culture and the establishment of cell lines
Introduction
Establishment of primary cultures from various sources
Source of material
Isolation of cells
Substrate for attachment
Culture conditions
Evolution of primary cultures
Nomenclature
Subculture
Growth phases
Characteristics of limited lifespan cultures
Lifespan and senescence
Phenotype
Karyotype
Establishment of continuous cell lines
Spontaneous
Chemical transformation
Viral transformation
Properties of continuous cell lines
Aneuploidy
Heterogeneity and instability
Differentiated status
Tumorigenicity
Cell fusion
Methods
Properties of hybrids
Applications
Genetic engineering techniques
Introduction of genes
Oncogene immortalization
Safety considerations
Concluding remarks
Advantages of cell culture over in vivo experimentation
Limitations
Acknowledgements
References
Specific cell types and their requirements
Introduction
General principles
Growth requirements of cells in vitro
Epithelial cells
Mesenchymal cells
Neuro-ectodermal cells
Gonadal cells
Embryonal stem (ES) cells
Leukocytes
Insect cells
Conclusion
Acknowledgements
References
Cloning
Introduction
Development of techniques
Uses of cloning
Limitations of cloning
Special requirements of cells growing at very low densities
Cell cloning procedures
Choice of technique
Methods applicable to both attached and suspension cells
Methods for attached cells
Methods for suspension cells
References
The quality control of cell lines and the prevention, detection, and cure of contamination
Introduction
Obtaining the basic material
Importance of cell culture collections
Resource centres
Cell culture databases
Quarantine and initial handling of cell lines
Accessioning scheme
Laboratory design
Handling of cell lines
Production of cell banks
Microbial quality control
Sources of microbial contamination
Testing for bacteria, yeasts, and other fungi
Testing for mycoplasma
Virus testing
Bovine spongiform encephalopathy
Elimination of contamination
Authentication
Isoenzyme analysis
Cytogenetic analysis
DNA fingerprinting
Regulatory aspects
References
Good Laboratory Practice in the cell culture laboratory
What happens if you fall under a bus?
GLP, an overview
What is GLP and when should it be used?
Planning the study
Performance of the study
Personnel
Methods
Equipment
Materials
Monitoring
Facilities and systems
Study-specific audits
Final report audit
Records
Advice for researchers creating new cell lines
Acknowledgements
References
List of suppliers
Culture collections and other resource centres
Online resources for cell biology
Index