Virus Culture A Practical Approach

by Cann, Alan J.

• ISBN13: 9780199637140
• ISBN10: 0199637148
• Edition: 1st
• Format: Paperback
• Copyright: 2000-05-11
• Publisher: Oxford University Press

Summary

Virus Culture: A Practical Approach provides a broad treatment of the principles and practice of virus culture and will be of interest to all those, whether in academic, industrial, or clinical research, involved in virus culture. The first chapter is an overview of cell culture techniques essential for virologists. Other techniques then covered are isolating, identifying, concentrating, and purifying viruses. Electron Microscopy as applied to virology is also explained. Chapter 6 is about creating virus vaccines and chapters 7 and 8 cover antiserum production, monoclonal antibodies and antiviral drug testing. The final chapter describes the methods used to study plant viruses.

Table of Contents

List of Contributors	xv
Abbreviations	xvii
Cell culture	1	(32)
K. Blake
A. Stacey
Introduction	1	(4)
Sources of cell cultures	1	(1)
Suppliers of cell cultures	2	(1)
Types of cell culture	3	(2)
Culture media	5	(2)
Basal media	5	(1)
Serum	6	(1)
Serum-free media	6	(1)
Antibiotics	7	(1)
Good practice in cell culture	7	(1)
Principles of cell banking	8	(1)
Subculture of cells	9	(2)
Quantification of cell cultures	11	(2)
Cryopreservation and storage of cell lines	13	(3)
Storage of cryopreserved cell stocks	15	(1)
Resuscitation of cryopreserved cell stocks	15	(1)
Quality control testing of cell lines	16	(14)
Preparation of cell cultures prior to testing	16	(1)
Mycoplasma	17	(7)
Bacteria and fungi	24	(2)
Viruses	26	(2)
Isoenzyme analysis	28	(1)
Karyotyping	29	(1)
DNA fingerprinting	29	(1)
Transportation	30	(3)
Shipment of growing cultures	30	(1)
Shipment of frozen materials on dry ice (solid carbon dioxide)	31	(1)
References	31	(2)
Virus isolation	33	(28)
A. J. Cann
W. Irving
Why culture viruses?	33	(2)
Viruses not amenable to cultivation	35	(6)
Treatment of clinical specimens for virus isolation	41	(4)
Identification of virus replication	45	(1)
Passaging of viruses	46	(2)
Confirmation of virus isolation	48	(4)
Adaptations of cell culture	52	(7)
Haemadsorption test	54	(1)
Detection of early antigen fluorescent foci (DEAFF test)	55	(4)
Conclusions	59	(2)
References	60	(1)
Concentration and purification of viruses	61	(20)
P. D. Minor
The need for virus concentration and purification	61	(3)
Practical considerations	62	(2)
Virus concentration: general principles	64	(3)
Purification of virus: identification of virus-containing fractions	67	(1)
Purification by ultracentrifugation	68	(10)
Types of ultracentrifugation gradient	68	(1)
Forming gradients	69	(9)
Assessment of purity	78	(3)
References	80	(1)
Assays for virus infection	81	(30)
J. J. Gray
Introduction	81	(1)
Identification of viruses isolated in cell culture	81	(7)
Cytopathic effects	81	(1)
Virus infectivity assays	81	(4)
Immunofluorescence	85	(1)
Electron microscopy	85	(1)
Other methods of identifying viruses growing in cell culture	86	(1)
Detection of virus pre-cytopathic effects	87	(1)
Detection of viruses not producing a cytopathic effect	87	(1)
Virus antigen detection	88	(3)
Immunofluorescence	88	(1)
Particle agglutination	89	(1)
Antigen-capture ELISA	90	(1)
Virus genome detection	91	(9)
Nucleic acid extraction	91	(2)
Genome detection assays	93	(6)
Quantitative nucleic acid amplification techniques	99	(1)
Detection of specific anti-viral antibodies	100	(11)
Virus-specific IgM detection	102	(2)
Detection of virus-specific IgG	104	(4)
References	108	(3)
Electron microscopy of viruses	111	(38)
H. R. Gelderblom
S. Biel
Introduction	111	(7)
Virus structure and strategies for virus detection	118	(1)
Specimen preparation	118	(5)
Centrifugation	121	(1)
Ultracentrifugation	121	(1)
Ultrafiltration	122	(1)
Precipitation	122	(1)
Bioaffinity techniques	122	(1)
EM grids and support films	123	(1)
Negative staining	124	(8)
Negative stains: properties	126	(2)
Negative staining procedures	128	(2)
Quantitative aspects: measuring and counting in TEM	130	(2)
Embedding and thin sectioning	132	(6)
Fixation and staining	132	(2)
Embedding	134	(1)
Thin sectioning	135	(1)
Post-staining	136	(2)
Immuno-electron microscopy (IEM)	138	(8)
Antibodies: properties and usage	139	(2)
Immuno-negative staining	141	(2)
Immuno-labelling of thin sections	143	(3)
Limitations of TEM in cell culture	146	(1)
Safety precautions	146	(3)
References	147	(2)
Virus vaccines	149	(34)
R. Jennings
C. W. Potter
Introduction	149	(3)
Non-living virus vaccines	152	(4)
Non-living vaccines produced by conventional methods	153	(2)
Non-living vaccines produced by recombinant DNA technology	155	(1)
Live, attenuated virus vaccines	156	(5)
Existing live attenuated vaccines	156	(4)
Live, attenuated vaccines based on reassortment	160	(1)
Delivery of vaccine antigens through vector technology and live, recombinant micro-organisms	161	(3)
DNA vaccines	164	(3)
Replication-deficient viruses as vaccines	167	(2)
Enhancement strategies for virus vaccines	169	(4)
In vitro and in vivo models in vaccine development	173	(2)
Control and licensing of virus vaccines	175	(4)
Future developments	179	(4)
References	181	(2)
Antiserum production and monoclonal antibodies	183	(18)
J. McKeating
C. Shotton
M. Valeri
Introduction	183	(1)
Antigen preparation	183	(2)
Synthetic peptides	184	(1)
Production of antisera	185	(2)
Immunization	185	(2)
Production of monoclonal antibodies	187	(12)
Immunization	188	(4)
Analysis of immune response	192	(3)
Hybridoma production	195	(3)
Analysis of fusions	198	(1)
Isotyping	199	(1)
Purification	199	(2)
References	200	(1)
Antiviral drug testing	201	(38)
J. S. Oxford
L. S. Kelly
S. Davies
R. Lambkin
Introduction	201	(2)
Inhibitors of critical viral enzymes	202	(1)
The role of cell culture in testing antiviral compounds	203	(2)
In vitro analysis	204	(1)
The design of the anti-viral testing laboratory	205	(4)
Testing procedures	209	(14)
In vitro testing of compounds for antiviral effect against Influenza A	211	(5)
In vitro testing of compounds for antiviral effect against herpes simplex	216	(3)
In vitro testing of compounds for antiviral effect against HIV-1	219	(3)
Medium to high throughput screening for potential antiviral compounds effective against HIV-1	222	(1)
Animal model infections	223	(9)
Influenza virus animal model systems	223	(5)
Mouse retrovirus model	228	(3)
Herpes simplex guinea pig model	231	(1)
Genetic analysis of virus resistance to antiviral drugs	232	(3)
Conclusions	235	(4)
References	237	(2)
Plant virus culture	239	(28)
E. P. Rybicki
S. Lennox
Introduction	239	(2)
Isolation of plant viruses	241	(11)
Virus isolation	241	(3)
Virus purification	244	(1)
Purification of virus genomic nucleic acid	245	(7)
Tissue culture and plant viruses	252	(8)
Callus initiation	253	(2)
Establishing a suspension culture	255	(2)
Production of somatic embryos from regenerable type II cultures	257	(1)
Transient virus replication in callus or suspension cells	257	(3)
Assay of plant viruses	260	(7)
PCR amplification	261	(1)
References	262	(5)
Appendix 1. Suppliers of specialist items	267	(4)
Index	271