Table Of ContentImmunochemistry
of the
Extracellular
Matrix
Volume II
Applications
Editor
Heinz Furthmayr, M.D.
Associate Professor
Department of Pathology
Yale University
New Haven, Conneticut
Boca Raton London New York
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Library of Congress Cataloging-in-Publication Data
Main entry under title:
Immunochemistry of the extracellular matrix.
Bibliography: p.
Includes index.
1. Collagen. 2. Immunochemistry.
3. Ground substance (Anatomy) 4. Extra-
cellular space. I. Furthmayr, Heinz.
II. Title: Extracellular matrix.
QP552.C6145 616.7’9 81-18062
ISBN 0-8493-6196-6 (v. 1) AACR2
ISBN 0-8493-6197-4 (v. 2)
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PREFACE
Fibrous connective tissue is the single most prevalent tissue in the body and in
a variety of forms, such as bone, tendon, cartilage, and fascia, it gives structural
rigidity. Collagen, as the major protein constituent of connective tissue is found in
addition throughout the body forming structures such as basement membranes, and
it is part of the extracellular matrix to which cells adhere to form specific organs.
Five genetically distinct types of collagen with distinct physicochemical and struc-
tural properties have been described thus far. These collagens are synthesized by
various mesenchymal and epithelial cells and apparently are distributed in tissues
and organs in specific ways. In a variety of pathological conditions expression,
distribution and control of production and/or degradation are affected.
The main purpose of the two volumes on Immunochemistry of the Extracellular
Matrix is to describe state of the art methods, which have been proven to provide
antibody reagents of defined specificity to collagens as well as other glycoproteins
found in association within connective tissue. The isolation and purification of col-
lagens, procollagens, and related connective tissue proteins are described for several
tissues, tissue culture cells and species. Immunization with these collagens in lab-
oratory animals yields antibodies with different characterization specificities: to the
procollagen extension fragments, the non-helical segments of the a-chains, to helical
and denatured determinants. Methods are described to obtain, isolate, and char-
acterize these antibody specificities by selection of the appropriate animal for im-
munization, immunoadsorbtion procedures, and sensitive serological assays. Ex-
perimental data are given and previous results are discussed in terms of producing
antibody reagents of desired specificity. Also included are immunohistological meth-
ods at the level of the right microscope and the electron microscope aimed at elu-
cidating the localization of these collagens in vivo in a variety of tissues.
Connective tissue biochemistry, biology, and pathology are rapidly expanding
fields today and the information assembled in these volumes should encourage and
facilitate the use of immunochemical tools for studies in these important areas of
research. Finally, the authors tried to define several challenging areas of inquiry in
connective tissue biology in which specific antibodies can contribute to our under-
standing of cellular processes.
The task of assembling these two volumes was greatly facilitated by the enthusiasm
of the contributing authors, all experts in the field, who regarded this project as a
worthwhile and timely effort and I am grateful for their support, effort, and valuable
contributions.
In writing my own contributions I came to realize more than ever, how much I
owe to my teachers Rupert Timpl and Klaus Kiihn, and to the many friends I have
found amongst the group of connective tissue biochemists. Amongst the individuals
at Yale I particularly wish to thank Joseph Roll and Joseph Madri, who not only
helped to bring this project to fruition, but who were also responsible for initiating
connective tissue reseach in the Department of Pathology together with me.
I welcome comments and criticisms from readers.
November 1981 Heinz Furthmayr
THE EDITOR
Heinz Furthmayr, M.D., University of Vienna, Austria, is Associate Professor of
Pathology at Yale University. He received his training in immunochemistry in the
Department of Immunochemistry in Munich, Germany, and in membrane biochem-
istry at the Department of Pathology at Yale University. He has been a Visiting
Fellow in the Department of Experimental Biology and Genetics at the CSSR Acad-
emy of Sciences in Prague and at the Hopital St. Louis in Paris. In 1978 he received
a Research Career Award from the American Cancer Society. His publications
include over 70 scientific articles, several reviews and chapters on connective tissue
proteins and plasma membrane glycoproteins. He is a member of the American
Society of Immunologists and the American Society of Pathologists and serves on
Advisory Committees of the U.S. Public Health Service.
CO NT RIB UTORS
Volume II
John Bateman, Ph.D. Klaus von der Mark, Ph.D.
Visiting Fellow Head of Research Group
National Cancer Institute Max-Planck-Institute for Biochemistry
National Institutes of Health Martinsreid, West Germany
Bethesda, Maryland
Bjorn R. Olsen, Ph.D.
Mario Chojkier, M.D. Professor of Biochemistry
Consultant College of Medicine and Dentistry
National Cancer Institute Rutgers Medical School
National Institutes of Health Piscataway, New Jersey
Bethesda, Maryland
Donna M. Pesciotta
Research Associate
Heinz Furthmayr, M.D.
Department of Anatomy
Associate Professor
Albert Einstein College of Medicine
Department of Pathology
Bronx, New York
Yale University
New Haven, Connecticut
Beverly Peterkofsky, Ph.D.
Research Biochemist
Gary R. Grotendorst, Ph.D.
National Cancer Institute
Postdoctoral Fellow
National Institutes of Health
National Institute of Dental Research
Bethesda, Maryland
National Institutes of Health
Bethesda, Maryland David H. Rohrbach, Ph.D.
Postdoctoral Fellow
A. Tyl Hewitt, Ph.D. National Institute of Dental Research
NIH Expert National Institutes of Health
National Institute of Dental Research Bethesda. Maryland
National Institutes of Health
F. Joseph Roll, M.D.
Bethesda, Maryland
Assistant Professor
Department of Medicine
John D. Kemp, M.D.
Liver Center
Postdoctoral Fellow in Immunology and
University of California
Pathology
San Francisco, California
Department of Pathology
Yale University H. E. J, Seppii, M.D.
New Haven, Connecticut Professor of Anatomy
Oulu Medical School
Hynda K. Kleinman, Ph.D. Oulu, Finland
Research Chemist
National Institute of Dental Research Elliott Schiffman, Ph.D.
National Institutes of Health Research Chemist
Bethesda, Maryland National Institute of Dental Research
National Institutes of Health
Joseph A. Madri, M.D., Ph.D. Bethesda, Maryland
Assistant Professor
Department of Pathology Victor P. Terranova, D.D.S., Ph.D.
Yale University Staff Fellow
New Haven, Connecticut National Institute of Dental Research
National Institutes of Health
George R. Martin, Ph.D. Bethesda, Maryland
Chief, Laboratory of Developmental Biology
and Anomalies Rupert Timpl, Ph.D.
National Institute of Dental Research Head of Research Group
National Institutes of Health Max-Planck-Institute for Biochemistry
Bethesda, Maryland Martinsreid, West Germany
Hugh H. Varner, Ph.D. Charlotte M. Wilkes
Postdoctoral Fellow Medical Student
National Institute of Dental Research Eastern V.A. Medical College
National Institutes of Health Virginia Beach. Virginia
Bethesda. Maryland
TABLE OF CONTENTS
Volume I
Chapter 1
Chemical Properties of Collagen
Klaus Kiihn
Chapter 2
Methods of Collagen Fractionation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3 1
Robert L. Trelstad
Chapter 3
Preparation of Type I and Type III Collagens, their PN Collagens and
Aminoterminal Extension Peptides . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 43
R. W. Glanville
Chapter 4
Preparation of Type II Collagen .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. 55
William T. Butler
Chapter 5
Isolation and Characterization of Pepsin Solubilized Basement Membrane
Collagens (Type IV) from Human Placenta, Bovine Kidney Cortices, and
Bovine Lens Capsule . . . . . . . . . . . .. . . . . . . . . . . . . .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 61
Saryu N. Dixit and R. W. Glanville
Chapter 6
The Preparation of Type V (AB Collagen .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. . 75
2)
Joseph A. Madri ·
Chapter 7
Preparation of Antisera Specific for the Amino and Carboxyl Propetides of
Type I and Type II Procollagen .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. 91
D. M. Pesciotta, S. Curran, and B. R. Olsen
Chapter 8
Isolation of Fibronectin from Plasma and Cells .. .. .. .. .. . .. .. .. .. .. .. .. .. .. .. .. .. .. .. . 11 1
Kenneth M. Yamada
Chapter 9
Laminin, a Glycoprotein from Basement Membranes ............................... 125
Jean-Michel Foidart, Rupert Timpl, Heinz Furthmayr, and George R. Martin
Chapter tO
Isolation and Properties of Chondronectin, the Chondrocyte Attachment
Factor .............................................................................................. 135
A. Tyl Hewitt, Hugh H. Varner, and George R. Martin
Chapter II
Immunization Procedures, Isolation by Affinity Chromatography and
Serological and Immunochemical Characterization of Collagen Specific
Antibodies . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 143
Heinz Furthmayr
Chapter 12
Production of Monoclonal Antibodies to Collagens and their
Immunofluorescence Localization in Embryonic Cornea and Cartilage . . . . . . . . 179
Thomas F. Linsenmayer and Mary J. C. Hendrix
Chapter 13
Radioimmunoassays in Studies of Connective Tissue Proteins . . . . . . . . . . . . . . . . . . . 199
Rupert Timpl and Leila Risteli
Chapter 14
Enzyme Linked Immunoassay (ELISA) for Connective Tissue Proteins:
Type I Collagen . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 237
Stephen I. Rennard, George R. Martin, and Ronald G. Crystal
Index 253
Volume II
Chapter I
The Cell Biology of Collagen Secretion
D. M. Pesciotta and B. R. Olsen
Chapter 2
Determination of Collagen Synthesis in Tissue and Cell Culture Systems ..... 19
Beverly Peterkofsky, Mario Chojkier, and John Bateman
Chapter 3
Immunocytochemical Techniques in Connective Tissue Research . . . . . . . . . . . . . . . 49
F. J. Roll and J. A. Mardri
Chapter 4
The Use of Antibodies to Connective Tissue Proteins in Studies on their
Localization in Tissues . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 89
Heinz Furthmayr and Klaus von der Mark
Chapter 5
Components of Basement Membranes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 119
Rupert Timpl
Chapter 6
Collagenous Matrices as Determinants of Cell Function . . . . . . . . . . . . . . . . . . . . . . . . . . . . 151
H. K. Kleinman, D. H. Rohrbach, V. P. Terranova, H. H. Varner, A. T.
Hewitt, G. R. Grotendorst, C. M. Wilkes, G. R. Martin, H. Seppa, and E.
Schiffm ann
Chapter 7
The lmmunobiology and Immunogenetics of the Collagens ........................ 175
John D. Kemp and Joseph A. Madri
Index ............................................................................................... 187
