Monoclonal Antibody and Immunosensor Technology: The Production and Application of Rodent and Human Monoclonal Antibodies

455 Pages·1991·17.28 MB·English

by  Ailsa M. Campbell (Eds.)

Monoclonal antibody
and immunosensor
technology
The production and
application of rodent
and human monoclonal antibodies
LABORATORY TECHNIQUES
IN BIOCHEMISTRY AND
MOLECULAR BIOLOGY
Volume 23
Edited by
P.C. van der VLIET - Department for Physiological Chemistry,
University of Utrecht, Utrecht
ELSEVIER
AMSTERDAM. LONDON. NEW YORK . TOKYO
MONOCLONAL ANTIBODY
AND IMMUNOSENSOR
TECHNOLOGY
The production and application of rodent
and human monoclonal antibodies
Ailsa M. Campbell
Department of Biochemistry,
University of Glasgow,
Glasgow GI2 SQQ,
U.K.
1991
ELSEVIER
AMSTERDAM. LONDON. NEW YORK . TOKYO
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Preface
This book is the successor to a book in the same series entitled Mono-
clonal Antibody Technology, published in 1984. Even at that time it
was apparent that unrealistic projections were being made for a useful
but not omnipotent technology. The main problem is, and has always
been, that it is possible to obtain invaluable monoclonal antibodies
as reagents, and also to obtain useless ones. The skill lies in the selec-
tion of high affinity Mabs by a method which is close to the final use
and far too many research strategies have concentrated on quantity
(numbers of positive clones) rather than quality. It has often been dis-
tressing to see the technique abused by those who do not appreciate
the importance of affinity and native antigen presentation.
The basic method for Mab production remains unchanged and fu-
sion and cloning methods are close to those used 10 years ago. The
main changes relate to the methodology of selection and use. In the
early 1980s people sought Mabs to simple soluble proteins and there
is now a vast library of Mabs to abundant proteins such as lysosyme,
all with no application. In contrast, there is an increasing interest in
Mabs to cell membrane proteins and these are scarcely ever produced
by the generation of Mabs to denatured proteins or peptides, since
epitopes are usually conformational. Thus selection techniques have
changed.
Too many researchers fail to understand that a monoclonal anti-
body is not necessarily monospecific. I have emphasised this point
heavily in various sections of the book. A good Mab is usually mono-
specific unless slammed up against the wall of an ELISA plate against
molar amounts of irrelevant antigen in which case it may give a slight
signal. A bad Mab will cross-react with a wide range of antigens. The
methods of obtaining the former are detailed.
V
vi
This book has a slight bias towards Mabs directed to supposed tu-
mour antigens. I have been astonished by the frequency with which
Mabs to uncharacterised antigens have been used, to predictably little
effect in ‘tumor therapy’, especially as the adverse side effects of a
mouse Mab are considerable (Section 1.9). Almost all these Mabs are
to high density structural antigens which, in the light of modern un-
derstanding of oncogene product function, are irrelevant to tumour
progression. This tendency is unfortunate, since far better Mabs could
be made in the light of modern knowledge of the nature of tumour
progression.
The book also covers the concepts behind the emerging biosensor
techniques (Chapter 12). Frankly, having made my way through the
mystiques of the systems, I was surprised by their primitive low detec-
tion limits despite their cost and cumbersome nature. It would appear
that there has been a breakdown in communications between Mab
technologists and electrochemical or opto-electronic engineers where
the latter group have assumed that antibodies have exquisite specifi-
city and no cross-reactive potential and the former have failed to dis-
abuse them of this fact. In terms of classic detection systems, for my
own work, I am now proceeding to use enzyme-linked chemilumines-
cent detection systems and all instant camera.
I have also covered the gene cloning strategies for Mabs and again
have found that many of the complex systems, once understood and
applied, are unsuitable for Mab production. In particular, PCR gene-
rated variable gene libraries (Section 9.6) are technically unsound for
reasons stated in the text. However, other aspects of genetic manipu-
lation involving PCR are very exciting and the prospect of immortal-
king human Mab responses by PCR of early unstable, but high affini-
ty clones seems to have immense potential. Tobacco plants and alfalfa
looping moths notwithstanding, the best expression system is still a
myeloma cell.
vii
I have included a section on Mab patents which I particularly en-
joyed investigating. The fact that the original Kohler-Milstein fusion
procedure was not patented means that no financial concern or com-
pany promotes it. I should like to do so. It is still the best way of mak-
ing a good Mab.
Ailsa Campbell
January 199 1.
This Page Intentionally Left Blank
Acknowledgements
I should like to thank Celia Cannon, Pat Ferry, Souravi Ghosh, Lor-
raine Haynes and Delyth Wong in particular.
I should also like to acknowledge help from Munir Alam, Peggy
Anderson, Rob Butcher, Flora Campbell, Magnus Campbell, Sue
Christie, Bill Cushley, Bertil Damato, Chris Darnborough, Morag
Davidson, Fiona Durie, Alan Hughes, Janet Jones, Moira McCann,
Tom Mathieson and Ian Ramsden.
ix