Table Of ContentMethods in
Mi c ro bio lo gy
Volume 25
Im m u n ology of Infection
Edited by
Stefan HE Kaufmann
Department of Immunology
University of Ulm
Germany
and
Dieter Kabelitz
Paul-Ehrlich Institute
Langen
Germany
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Contributors
Reinhard Andreesen Department of Hematology and Oncology, University of
Regensburg, D-94042 Regensburg, Germany
Ingo B Autenrieth Max von Pettenkofer Institute for Hygiene and Medical
Microbiology, Ludwig-Maximilians University Munich, Pettenkoferstrasse 9a,
D-80336 Munich, Germany
Jacques Banchereau Baylor Research Institute, 3535 Worth Street, Dallas, TX 75246,
USA
John T Belisle Mycobacteria Research Laboratories, Department of Microbiology,
Colorado State University, College of Veterinary Medicine, Fort-Collins, CO 80523,
USA
Jay A Berzofsky Molecular Immunogenetics and Vaccine Research Section, National
Cancer Institute, National Institutes of Health, Bldg 10, Rm 6B12, Bethesda, MD 20892,
USA
Willi Born Department of Medicine, National Jewish Center for Immunology and
Respiratory Medicine, 1400 Jackson Street, Denver, CO 80206, USA
James F Brown Department of Pathobiological Sciences, School of Veterinary
Medicine, 2015 Linden Drive West, Madison, WI 53706, USA
Kevin Brunson College of Business Administration, Room 315, University of Notre
Dame, Notre Dame, IN 46556, USA
Dirk H Busch Section of Infectious Diseases, Yale University School of Medicine, 803
LCI, New Haven, CT 06520, USA
Carol Cady Department of Medicine, National Jewish Center for Immunology and
Respiratory Medicine, 1400 Jackson Street, Denver, CO 80206, USA
Christophe Caux Schering-Plough, Laboratory for Immunological Research, 27
Chemin des Peupliers, BP 11,69571 Dardilly, France
Andrea M Cooper Mycobacteria Research Laboratories, Department of Microbiology,
Colorado State University, College of Veterinary Medicine, Fort-Collins, CO 80523,
USA
Charles J Czuprynski Department of Pathobiological Sciences, School of Veterinary
Medicine, 2015 Linden Drive West, Madison, WI 53706, USA
Frangoise Cottrez INSERM U343, HBpital de l’Archet, Route de Saint Antoine de
Ginestiere, 06202 Nice Cedex 3, France
vii
Anne S De Groot TB/HIV Research Laboratory, International Health Institute, Brown
University School of Medicine, Providence, RI 02912, USA
Colette Dezutter-Dambuyant INSERM Urn, Hepita1 Edouard Herriot, 69374 Lyon,
France
John J Donnelly Department of Virus and Cell Biology, Merck Research Laboratories,
WP16-3, West Point, PA 19486, USA
Rainer Duchmann 1 Medizinische Klinik und Poliklinik, Johannes Gutenberg
Universitat, LangenbeckstraBe 1,55101 Mainz, Germany
Stefan Ehlers Division of Molecular Infection Biology, Research Center Borstel,
Pakallee 22,23845 Borstel, Germany
Robert Etges University of Notre Dame, Department of Biological Sciences, 214
Galvin Life Science, Notre Dame, IN 46556, USA
Helmut Fickenscher Institut fiir Klinische und Molekulare Virologie, SchloBgarten 4,
D-91054 Erlangen, Germany
Bernhard Fleckenstein Institut fiir Klinische und Molekulare Virologie, SchloBgarten
4, D-91054 Erlangen, Germany
Kohtaro Fujihashi Immunobiology Vaccine Center and Departments of Oral Biology
and Microbiology, The University ,of Alabama at Birmingham, Birmingham, AL
352942170, USA
Johannes Gerdes Division of Molecular Immunology, Forschungzentrum Borstel,
Parkallee 22,23845 Borstel, Germany
Siamon Gordon Sir William Dunn School of Pathology, University of Oxford, South
Parks Road, Oxford OX1 3RE, UK
Richard Haworth Sir William Dunn School of Pathology, University of Oxford, South
Parks Road, Oxford OX1 3RE, UK
Hans J Hedrich Institut fiir Versuchstierkunde, Medizinische Hochschule Hannover,
30625 Hannover, Germany
Thomas K Held Department of Hematology and Oncology, Virchow Hospital,
Humboldt University, Berlin, Germany
Takachika Hiroi Department of Mucosal Immunology, Research Institute for
Microbial Diseases, Osaka University, Suita, Osaka 565, Japan
Bill M Jesdale TB/HIV Research Laboratory, International Health Institute, Brown
University School of Medicine, Providence, RI 02912, USA
viii
Dieter Kabelitz Department of Immunology, Paul-Ehrlich Institute, Postfach D-63207
Langen, Germany
Stefan Kaufmann Max-Planck-Institute for Infection Biology, Monbijoustr-2, D-10117
Berlin, Germany
Hiroshi Kiyono Department of Mucosal Immunology, Research Institute for
Microbial Diseases, Osaka University, Suita, Osaka 565, Japan
Stefan W Krause Department of Hematology and Oncology, University of
Regensburg, D-94042 Regensburg, Germany
Marina Kreutz Department of Hematology and Oncology, University of Regensburg,
D-94042 Regensburg, Germany
Pascale Kropf Imperial College School of Medicine at St Mary’s, Department of
Immunology, Norfolk Place, London W2 lPG, UK
Michael Lahn Department of Medicine, National Jewish Center for Immunology and
Respiratory Medicine, 1400 Jackson Street, Denver, CO 80206, USA
Margaret A Liu Department of Virus and Cell Biology, Merck Research Laboratories,
WP16-3, West Point, PA 19486, USA
Yong-Jun Liu DNAX, 901 California Avenue, Palo Alto, CA 94304, USA
Karin Lovgren-Bengtsson Swedish University of Agricultural Sciences, Faculty of
Veterinary Medicine, Department of Veterinary Microbiology, Section of Virology, Box
585, BMC, S-75123 Uppsala, Sweden
Erminia Mariani Istituto di Ricerca Codivilla Putti, Laboratorio di Immunologia e
Genetica, Via di Barbino 1/10, 1-40136 Bologna, Italy
Elisabeth Mirker-Hermann 1. Medizinische Klinik und Poliklinik, Johannes
Gutenberg Universitat, LangenbeckstraBe 1,55101 Mainz, Germany
Jerry R McGhee Immunobiology Vaccine Center and Departments of Oral
Biology and Microbiology, The University of Alabama at Birmingham, AL 35294-217 0,
USA
Cornelius JM Melief Department of Immunohaematology and Blood Bank,
University Hospital Building 1 E3-Q, NL-2230 RC Leiden, The Netherlands
Martin EA Mielke Institut fiir Medizinishe Mikrobiologie und Infektious-
immunologie, Freie Universitat Berlin, Hindenburgdamm 27,12203 Berlin, Germany
Donna L Montgomery Department of Virus and Cell Biology, Merck Research
Laboratories, WP16-3, West Point, PA 19486, USA
ix
Horst Mossmann Max-Planck-Institut fiir Immunbiologie, Stiibeweg 51, 79108
Freiburg, Germany
Akiko Mukasa Department of Medicine, National Jewish Center for Immunology and
Respiratory Medicine, 1400 Jackson Street, Denver, CO 80206, USA
Ingrid Muller Imperial College School of Medicine at St Mary’s, Department of
Immunology, Norfolk Place, London W2 IPG, UK
Werner Nicklas Deutsches Krebsforschungszentrm, Im Neuenheimer Feld 280,
69120 Heidelberg, Germany
Rebecca O’Brien Department of Immunology, University of Colorado, Health
Sciences Center, 4200 E 9th Avenue, Denver, CO 80262, USA
Rienk Offringa Department of Immunohaematology and Blood Bank, University
Hospital Building 1 E3-Q, NL-2230 RC Leiden, The Netherlands
Ian M Orme Mycobacteria Research Laboratories, Department of Microbiology,
Colorado State University, College of Veterinary Medicine, Fort-Collins, CO 80523,
USA
Eric Pamer Section of Infectious Diseases, Yale University School of Medicine, 803
LCI, New Haven, CT 06520, USA
Graham Pawelec Section for Transplantation Immunology and Immunohaematology,
Medizinische Universitatsklinik, D-72076 Tubingen, Germany
Klaus Pechhold Section of Endocrinology, Department of Medicine, University of
Ulm, Robert-Koch-Str. 8, D-89081 Ulm, Germany
Andreas Radbruch Deutsches Rheumaforschungszentm, Hannoverschestrai3e 27,
D-10115 Berlin, Germany
Christopher Reardon Department of Dermatology, University of Colorado Health
Sciences Center, Denver, CO 80262, USA
Alan D Roberts Mycobacteria Research Laboratories, Department of Microbiology,
Colorado State University, College of Veterinary Medicine, Fort-Collins, CO 80523,
USA
Alexander Scheffold Deutsches Rheumaforschungszentm, Hannoverschestrage 27,
D-10115 Berlin, Germany
Ulrike Seitzer Division of Molecular Immunology, Research Center Borstel, Parkallee
22,23845 Borstel, Germany
Rafael Solana SeMco de Immunologia, Hospital Universitario Reina Sofia, Av.
Menendez Pidal s/n, E-14004 Cbrdoba, Spain
X
Moriya Tsuji Department of Medical and Molecular Parasitology, New York University
School of Medicine, 341 East 25th Street, New York, NY 10010, USA
Jeffrey B Ulmer Department of Virus and Cell Biology, Merck Research Laboratories,
WP16-3, West Point, PA, 19486, USA
John L VanCott Immunobiology Vaccine Center and Departments of Oral Biology and
Microbiology, The University of Alabama at Birmingham, Birmingham, AL 35294-2170,
USA
Hans Yssel INSERM U454, Jmmunopathologie de l'asthme, HGpital Arnaud de
Villeneuve, 371 Avenue Doyen Gaston Giraud, 34295 Montpellier Cedex 05, France
xi
It all began in a highly intertwined way. The golden age of medical micro-
biology, which had its height just before the turn of the 19th century not
only witnessed the discovery of numerous medically important pathogens,
but was also the cradle of immunological research. R. Koch discovered
Bacillus anthracis and Mycobacterium tuberculosis, and was also the first to
describe delayed type hypersensitivity reactions to bacterial products. The
work of L. Pasteur was both instrumental in establishing germ theory, and
also laid the basis for rational vaccine development. E. Behring and S.
Kitasato were not satisfied by their success in obtaining pure cultures of
Clostridium tetani, and went on to develop vaccines against tetanus.
Similarly, the seminal discoveries by E. Metchnikoff of phagocytosis and
by J. Bordet of complement-mediated lysis, and the development of the
side-chain theory by P. Ehrlich, to name but a few, were at the very inter-
face between bacteriology and immunology. Thereafter, however, medical
microbiology and immunology went their own ways.
After a period of quiescence, medical microbiology has been revital-
ized by its amalgamation with molecular genetics and cell biology, to
embark towards an understanding of the molecular cross-talk between
the host cell and the microbial pathogen. Immunology dramatically
broadened its scope from pure antimicrobial defence towards general
topics concerned with its role in homeostasis and pathology within the
mammalian organism. Today, the majority of immunological sub-
disciplines have become virtually independent of their roots. Because of
its broad scope, immunology has had to develop various methodologies
of its own in addition to assimiIating strategies from other fields.
Immunology has been at the forefront in the development of several
techniques of general importance. With the discovery of monoclonal anti-
body techniques, ELISA and ELISPOT assays for the detection of
molecules and fluorescence-activated cell sorting (FACS) systems for the
detection and separation of cells have gained general importance. Equally
important is the assimilation of cell biology and molecular genetic
techniques. Therefore, it is hardly possible for immunologists to master
such a broad array of methodologies.
The need for comprehensive manuals on immunological methods has
been fulfilled by several large volumes, including the Immunology Methods
Manual (I. Lefkovits, Academic Press, London, 1997) and Current Protocols
in Immunology 0. E. Coligan, A. Kruisbeek, D. H. Margulies, E. M. Shevach
and W. Strober, Wiley, Chichester, 1995). These are complemented by
more specific books for immunologists and for scientists working in
related fields, such as Antibodies - A Laborato y Manual (E. Harlow and D.
Lane, Cold Spring Harbor Laboratory Press, New York, 1988), the
Laboratory Manual on Manipulating the Mouse Embryo (B. Hogan, R.
Beddington, F. Constantini and E. Lacy, Cold Spring Harbor Laboratory
Press, New York, 1994) and, last not least, the famous standard work on
Molecular Cloning 0. Sambrook, E. F. Fritsch and T. Maniatis, Cold Spring
xiii
Harbor Laboratory Press, New York, 2nd edition, 1989). However, whilst
being invaluable for immunologic laboratories, these manuals are often
too broad for the microbiologist interested in using immunological
methods to analyse host-pathogen relationships. It is these scientists for
whom the present manual has been devised.
Accordingly, in preparing this volume, emphasis was placed on
selection with the risk of bias. Sophisticated immunological techniques, as
well as those mostly used in unrelated fields, such as tumour immunology
or transplantation immunology, were omitted on purpose. Similarly,
general techniques stemming from other fields, but now used widely in
immunology, were omitted because excellent manuals are already avail-
able, and the reader is referred to those. These include general molecular
cloning and cell biology techniques, as well as immunological techniques
of broader application, such as the generation of monoclonal antibodies.
On which methodologies then did we focus? First, emphasis was placed
on immune responses to bacteria, although many of the techniques
described are equally applicable to antiviral and antiprotozoal immunity.
Secondly, focus has been directed towards T cells, macrophages and
cytokines. The T lymphocyte is the central regulator of the anti-infective
immune response and the enormous increase in our knowledge about this
cell has only been made possible by the recent development of appropriate
techniques. T lymphocytes rarely combat infectious agents directly, but
rather do so with the help of macrophages and the dialogue between these
cells is mediated by cytokines. Equally large space, therefore, has been
reserved for the characterization of cytokines and professional phagocytes.
Although in vitro analyses are indispensible in investigating the immune
response against infectious agents, in vivo studies remain of critical
importance. In particular, evaluation of immunization strategies (such as
vaccination) is impossible without appropriate in vivo models. With the
advent of transgenic and gene deletion mouse mutants, novel strategies for
characterizing the role of defined molecules and cells in the in vivo setting
became feasible. Therefore, we considered it essential to include chapters
dealing with various aspects of experimental animal models.
We hope that this manual fills the gap between immunology and
microbiology and helps to re-establish a closer relationship between the
two disciplines. After all, infection is the outcome of the cross-talk
between prokaryotic and eukaryotic cells, and in mammals the immune
system has been given the task of being the major player.
We wish to cordially thank the care given by the editorial staff at
Academic Press, particularly Tessa Picknett and Duncan Fatz, as well as
our secretaries, Rita Mahmoudi and Constanze Taylor, for their great
dedication. Last, but not least, we are grateful to all our colleagues who,
by contributing to this manual, have generously let us share their extra-
ordinary expertise.
Stefan H. E. Kaufmann
Max-Planck-Institute for Infection Biology, Berlin, Germany
Dieter Kabelitz
Paul-Ehrlich Institute, Langen, Germany
xiv
Introduction: The immune
response to infectious agents
SHE Kaufmann
Department of Immunology, University of Ulm, and
Max-Planck-Institute for Infection Biology, Berlin, Germany
D Kabelitz
PauCEhrlich Institute, Langen, Germany
CONTENTS
Introduction
The innate immune system
The adaptive immune system
Cytokines
Concluding remarks
++++++
INTRODUCTION
It is the task of the immune system to protect the host against invading
infectious agents and thereby to prevent infectious disease. A plethora of
microbial pathogens exists (i.e. viruses, bacteria, fungi, parasites and
helminths) that have exploited strategies to circumvent an attack by the
immune system. Conversely, the immune system has evolved to provide
appropriate defence mechanisms at various levels of ‘non-specific’
(innate) and ‘specific‘ (adaptive) immune responses. In many instances,
an appropriate immune response to an infectious agent requires recipro-
cal interactions between components of the innate and the adaptive
immune systems.
The various micro-organisms have developed different strategies to
invade their host. Viruses make use of the host cell’s machinery for repli-
cation and are thus intracellular pathogens. Helminths, the other extreme,
are large organisms that cannot live within host cells, but rather behave as
extracellular pathogens. In between are bacteria, which, depending on the
species, live within or outside host cells, and protozoa where the extracel-
lular or intracellular localization may depend on the stage of their life cycle.
METHODS IN MICROBIOLOGY, VOLUME 25 Copyright 0 1998 Academic Press Ltd
ISBN 0-12-52152c2 All rights of reproduction in any form reserved
