Virus culture : a practical approach

291 Pages·1999·17.812 MB·English

by  Alan Cann

Virus Culture
The Practical Approach Series
SERIES EDITOR
B. D. HAMES
School of Biochemistry and Molecular Biology
University of Leeds, Leeds LS2 9JT, UK
See also the Practical Approach web site at http://www.oup.co.uk/PAS
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Affinity Chromatography if Cell Separation
Affinity Separations Cellular Calcium
Anaerobic Microbiology Cellular Interactions in
Development
Animal Cell Culture
(2nd edition) Cellular Neurobiology
Animal Virus Pathogenesis * Chromatin
Antibodies I and II * Chromosome Structural
Analysis
Antibody Engineering
Clinical Immunology
Antisense Technology
Complement
Applied Microbial
Physiology if Crystallization of Nucleic
Acids and Proteins
Basic Cell Culture
(2nd edition)
Behavioural Neuroscience
Cytokines (2nd edition)
Bioenergetics
The Cytoskeleton
Biological Data Analysis
Diagnostic Molecular
Biomechanics - Materials
Pathology I and II
Biomechanics — Structures and
DNA and Protein Sequence
Systems
Analysis
Biosensors
DNA Cloning 1: Core
Carbohydrate Analysis Techniques (2nd edition)
(2nd edition)
DNA Cloning 2: Expression
Cell-Cell Interactions Systems (2nd edition)
The Cell Cycle DNA Cloning 3: Complex
Cell Growth and Apoptosis Genomes (2nd edition)
DNA Cloning 4: Mammalian * HPLC of Macromolecules
Systems (2nd edition) (2nd edition)
* DNA Microarrays Human Cytogenetics I and II
(2nd edition)
* Drosophila (2nd edition)
Human Genetic Disease
Electron Microscopy in
Analysis
Biology
* Immobilized Biomolecules in
Electron Microscopy in
Analysis
Molecular Biology
Immunochemistry 1
Electrophysiology
Immunochemistry 2
Enzyme Assays
Immunocytochemistry
Epithelial Cell Culture
* In Situ Hybridization
Essential Developmental
(2nd edition)
Biology
Iodinated Density Gradient
Essential Molecular Biology I
Media
and I
Ion Channels
* Eukaryotic DNA Replication
* Light Microscopy (2nd edition)
Experimental Neuroanatomy
Lipid Modification of Proteins
Extracellular Matrix
Lipoprotein Analysis
Flow Cytometry (2nd edition)
Liposomes
Free Radicals
Mammalian Cell
Gas Chromatography
Biotechnology
Gel Electrophoresis of Nucleic
Medical Parasitology
Acids (2nd edition)
Medical Virology
* Gel Electrophoresis of Proteins
* MHC Volumes 1 and 2
(3rd edition)
* Molecular Genetic Analysis of
Gene Probes 1 and 2
Populations (2nd edition)
Gene Targeting
Molecular Genetics of Yeast
Gene Transcription
Molecular Imaging in
if Genome Mapping Neuroscience
Glycobiology Molecular Neurobiology
if Growth Factors and Receptors Molecular Plant Pathology I
Haemopoiesis and II
if High Resolution Molecular Virology
Chromotography Monitoring Neuronal Activity
Histocompatibility Testing Mutagenicity Testing
HIV Volumes 1 and 2 * Mutation Detection
Neural Cell Culture Preparative Centrifugation
Neural Transplantation Protein Blotting
Neurochemistry (2nd edition) if Protein Expression
Neuronal Cell Lines Protein Engineering
NMR of Biological Protein Function (2nd edition)
Macromolecules
Protein Phosphorylation
Non-isotopic Methods in
Protein Purification
Molecular Biology
Applications
Nucleic Acid Hybridisation
Protein Purification Methods
Oligonucleotides and
Protein Sequencing
Analogues
Protein Structure (2nd edition)
Oligonucleotide Synthesis
Protein Structure Prediction
PCR 1
Protein Targeting
PCR 2
Proteolytic Enzymes
*PCR3:PCR In Situ
Hybridization Pulsed Field Gel
Electrophoresis
Peptide Antigens
RNA Processing I and II
Photosynthesis: Energy
Transduction * RNA-Protein Interactions
Plant Cell Biology Signalling by Inositides
Plant Cell Culture if Steroid/Nuclear Receptor
(2nd edition) Superfamily
Plant Molecular Biology Subcellular Fractionation
Plasmids (2nd edition) Signal Transduction
Platelets * Transcription Factors
(2nd edition)
Postimplantation Mammalian
Embryos Tumour Immunobiology
* Post-translational Modification * Virus Culture
Virus Culture
A Practical Approach
Edited by
ALAN J. CANN
Department of Microbiology and Immunology,
University of Leicester
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OXFORD
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Phreface
The properties of viruses are distinct from those of living organisms, which
makes the study of virology different from other areas of biology. Many
specialized techniques have been developed to study viruses, and yet these
have almost invariably found their way into mainstream biology. It is not
possible to define in a single volume the techniques of virology, but this volume
and its companions set out to illustrate the major experimental methods
currently employed by virologists.
When I was first approached by David Hames to edit this volume, my initial
reaction was to decline. However, on brief reflection I decided that the Practical
Approach series was important enough to the research community that I
should take my turn as an editor. Having agreed to edit Virology: a practical
approach, it was only during the course of our subsequent discussions that the
magnitude of what I had acquiesced in fully dawned on me. My thanks go to
David Hames (joint series editor) for his guidance in shaping this and the
accompanying volumes. Inevitably there will be those who feel that this or that
should have been included or left out. It is not possible to include everything
within the format of this series!
Most importantly, thanks must go to the contributors who were prepared to
share their combined expertise with the wider research community. Chapter 1
provides an overview of cell culture, essential for all virologists, and frequently
forgotten by those with a tendency to slip into bad habits. Chapters 2 and 4 are
complementary insights into the fundamentals of virology: isolating and identify-
ing viruses. Chapter 3 describes methods for concentrating and purifying viruses,
Chapter 5 is on electron microscopy as applied to virology, and Chapter 7 de-
scribes antiserum and monoclonal antibody production. Chapter 6 is unusual
for a Practical Approach volume in that it is rather more theoretical that most.
The reason for this is that it is difficult to cover all approaches to virus vaccines
in detail, but my feeling is that this is such a significant area of virology that a
reasonably comprehensive treatment deserved to be included. This chapter is
complemented by the methods for antiviral drug testing described in Chapter 8.
Finally, the last chapter describes the specialized techniques used to study
plant viruses, which will not be of direct relevance to all readers, but is import-
ant to an overall understanding of practical virology, and the impact that
molecular biology has had on this field.
Finally, a word of advice to students (and others!) who experience difficulties
with any of the protocols. If you cannot get a protocol you know to have
worked previously to work for you, there are two choices. Either troubleshoot
the whole protocol in a step-by-step fashion with reference to appropriate con-
trols, in order to identify and rectify problems, or use a different protocol. In
my experience, many experimental problems arise from mutilated protocols.
Preface
These procedures are sequential and cumulative, where each step is depend-
ent on the previous ones. It is not a good idea to insert steps from one protocol
into another, or to omit steps, until you have already performed the procedure
successfully several times. That said, all of the protocols in this volume are
tried and tested.
Leicester A. J. C.
May 1999
viii
Contents
List of Contributors xv
Abbreviations xvii
1. Cell culture 1
K. Blake and A. Stacey
1. Introduction 1
Sources of cell cultures 1
Suppliers of cell cultures 2
Types of cell culture 3
2. Culture media 5
Basal media 5
Serum 6
Serum-free media 6
Antibiotics 7
3. Good practice in cell culture 7
4. Principles of cell banking 8
5. Subculture of cells 9
6. Quantification of cell cultures 11
7. Cryopreservation and storage of cell lines 13
Storage of cryopreserved cell stocks 15
Resuscitation of cryopreserved cell stocks 15
8. Quality control testing of cell lines 16
Preparation of cell cultures prior to testing 16
Mycoplasma 17
Bacteria and fungi 24
Viruses • 26
Isoenzyme analysis 28
Karyotyping 29
DNA fingerprinting 29
9. Transportation 30
Shipment of growing cultures 30
Shipment of frozen materials on dry ice (solid carbon dioxide) 31
References 31