Table Of ContentSTM and SFM
in Biology
Edited by
Othmar Marti
Department of Physics
University of Konstanz
Konstanz, Germany
Matthias Amrein
Institute for Medical Physics and Biophysics
University of Miinster
Miinster, Germany
ACADEMIC PRESS, INC.
A Division of Harcourt Brace & Company
San Diego New York Boston London Sydney Tokyo Toronto
Cover photographs: (Left) 4'->i-Octyl-4-cyanobiphenyl (8CB) adsorbed on graphite. (Adapted from
Figure 4.2; courtesy of D. P. E. Smith.) (Top) Surface of a diatom taken from a pond at the Univer
sity of Konstanz, Germany. Image by SFM. (Courtesy of Achim Linder.) (Right) STM image in
surface view representation of freeze-dried recA-DNA complexes coated with Pt-Ir-C. (Adapted
from Figure 2.18; courtesy of Matthias Amrein.)
This book is printed on acid-free paper. @
Copyright © 1993 by ACADEMIC PRESS, INC.
All Rights Reserved.
No part of this publication may be reproduced or transmitted in any form or by any
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United Kingdom Edition published by
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Library of Congress Cataloging-in-Publication Data
STM and SFM in biology / edited by Othmar Marti, Matthias Amrein.
p. cm.
Includes bibliographical references and index.
ISBN 0-12-474500-8
1. Scanning tunneling microscopy. 2. Scanning force microscopy.
I. Marti, Othmar. II. Amrein, Matthias.
QH212. S35S76 1993
578'45--dc20 92-43104
CIP
PRINTED IN THE UNITED STATES OF AMERICA
93 94 95 96 97 98 BB 9 8 7 6 5 4 3 2 1
CONTRIBUTORS
Numbers in parentheses indicate the pages on which the authors' contributions begin.
M. Amrein (127), Institute for Medical Physics and Biophysics, University of
Munster, 4400 Munster, Germany
Hans-Jurgen Apell (275), Department of Biology, University of Konstanz,
D-7750 Konstanz, Germany
Patricia G. Arscott (259), Department of Biochemistry, University of
Minnesota, St. Paul, Minnesota 55108
G. Binnig (309), IBM Research Division, IBM Physics Group Munich, 8000
Munich 40, Germany
Victor A. Bloomfield (259), Department of Biochemistry, University of
Minnesota, St. Paul, Minnesota 55108
Jaime Colchero (275), Department of Physics, University of Konstanz,
D-7750 Konstanz, Germany
C. P. Czerny (309), Tierarztliche Fakultat der Ludwig-Maximilians-
Universitat Munich, 8000 Munich 22, Germany
J. E. Frommer (205), IBM Almaden Research Center, San Jose, California
95120, and Institut fur Physik der Universitat, CH-4056 Basel, Switzerland
H. Gross (127), Institute for Cell Biology, Swiss Federal Institute of
Technology, ETH-Honggerberg, 8092 Zurich, Switzerland
R. Guckenberger (127), MPI for Biochemistry, 8033 Martinsried, Germany
W. Haberle (309), IBM Research Division, IBM Physics Group Munich, 8000
Munich 40, Germany
J. Κ. H. Horber (309), EMBL, 6900 Heidelberg, Germany
H. G. Liebich (309), Tierarztliche Fakultat der Ludwig-Maximilians-
Universitat Munchen, 8000 Munchen 22, Germany | jx
χ Contributors
Achim Under (275), Department of Biology, University of Konstanz, D-7750
Konstanz, Germany
S. M. Lindsay (229), Department of Physics, Arizona State University, Tempe, Arizona
85287
Othmar Marti (1, 275), Department of Physics, University of Konstanz, D-7750
Konstanz, Germany
T. J. McMaster (177), Η. H. Wills Physics Laboratory, University of Bristol, Bristol BS8
1TL, United Kingdom
M. J. Miles (177), Η. H. Wills Physics Laboratory, University of Bristol, Bristol BS8
1TL, United Kingdom
F. Ohnesorge (309), IBM Research Division, Physics Group Munich, 8000 Munich 40,
Germany
D. P. E. Smith (205), IBM Physics Group Munich, 8000 Munich 40, Germany
N. J. Tao (229), Department of Physics, Florida International University, Miami, Florida
33199
Preface
The scanning tunneling microscope (STM) is the first of a new genera
tion of microscopes in which various physical quantities of surfaces are
locally probed by tiny sensors scanning in close proximity over them.
The STM and its first offspring, the scanning force microscope (SFM),
may resolve surface topography, as well as some electronic and me
chanical properties at the atomic scale, by combining a highly precise
scanning mechanism, a small probe size, and a localized site for prob
ing surface properties. These microscopes attracted the immediate
attention of the biological community, not least because they usually
perform equally well in ultrahigh vacuum, in ambient atmosphere, and
even in aqueous solution, the natural environment of most biological
macromolecules.
The chapters in this book discuss in detail the application of STM
and SFM in biology. The book is addressed to everyone who wants to
become acquainted with the applications of scanning probe techniques
in a wide field ranging from the most simple organic molecules, beau
tifully arranged in crystalline films, to dynamic events on the outer
membranes of living cells in their natural environments.
In the Introduction to this book, the fundamentals of STM, SFM,
and scanning probe microscopy (SXM), in general, are presented, and
major aspects of the instrumental designs are discussed. Researchers
who intend to design their own experimental setups will find this
section especially valuable. The extended bibliographies guide the
reader to the source publications and to further readings on every
subject.
In the next parts of this book, biological applications of STM and
SFM are presented. Each self-contained chapter is a review of a spe
cific topic by scientists active in research on the subject of their
contribution. This book covers most aspects of the field with respect
to both the investigated specimens and the experimental techniques
presented. The experiments are described such that they can be repro-
xi
xii Preface
duced in an appropriate environment. Some views expressed in the book are
controversial, and sometimes different authors express mutually incompatible
interpretations. The reader should accept this as an inevitable side effect of any
quickly evolving subject. While there have been a few books on SXM in general,
this is the first to be dedicated fully to biological applications.
Othmar Marti
Matthias Amrein
CHAPTER 1
SXM: An Introduction
Othmar Marti
Department of Physics
University of Konstanz
Konstanz, Germany
1.1 Overview
1.2 STM
1.2.1 Theory of the STM
1.2.2 The STM
1.2.3 Image Processing: An Introduction
1.2.4 Selected Experiments
1.2.5 Related Techniques
1.2.6 Additional References
1.3 SFM
1.3.1 Theory of Force Microscopy
1.3.2 How to Measure Small Forces
1.3.3 The Force Microscope
1.3.4 Selected Experiments
1.4 SNOM
1.5 Electrochemistry with SXM
1.6 Local Experiments
1.6.1 Light Mixing
1.6.2 Additional References
1.7 New Developments
1.7.1 Miniaturization of the STM
1.7.2 Other SXM
Appendix A: Background Plane Removal
Appendix B: Correction of Linear Distortions in Two and Three Dimensions
References
STM and SFM in Biology
Copyright © 1993 by Academic Press, Inc. Al rlights of reproduction in any form reserved. 1
2 Othmar Marti
1.1 OVERVIEW
All scanning probe microscopes (SXM) are based on similar principles. The aim
of this chapter is to discuss their common aspects, to elucidate their differences,
and to point to their possible applications in the field of biology. The scanning
tunneling microscope (STM), invented by Binnig and Rohrer (1982), serves as a
model system. A summary of the theory of the STM points out the different
operating modes and techniques, deals with the problem of imaging, and gives
resolution criteria.
A detailed introduction to the mechanical and electronic design of the STM
is presented. Design rules are worked out to help the builders of an STM and to
allow the users to judge their instruments. Important parts of any STM experi
ment are the data acquisition and the image processing. Critical points in the data
acquisition systems and common image processing techniques are worked out.
All the technical issues of the STM are equally valid for other SXM techniques.
The section on the STM concludes with the description of a few experi
ments. The application of the STM to the imaging of biological and organic
matter is treated in depth in other chapters of this book.
The scanning force microscope (SFM) is the most successful offspring of the
STM. The design principles worked out for the STM are equally valid for the
SFM. The additional critical points of an SFM are treated. Special emphasis is
given to the description of the various interaction forces and the force-sensing
techniques, including the scanning force and friction microscope (SFFM). The
section on the SFM is closed with the discussion of a few representative experi
ments. Again, Chapter 7 in this volume presents a review of the application of
SFM to biology.
To conclude, a summary of other scanning probe techniques is given,
together with an outlook. The interested reader might also want to consult
review articles on scanning probe microscopy, such as those by Binnig and
Rohrer (1986), Wickramasinghe (1989), Tersoff (1990b), and Rugar and Hansma
(1990) and books such as that by Sarid (1991), or the series edited by Guntherodt
and Wiesendanger (1992).
1.2 STM
The following sections give an overview of the theory of the STM, of design
principles commonly used to build SXM, and of image processing techniques
used in the field. Discussions of selected experiments and some related tech
niques complete the sections.
1.2.1 Theory of the STM
Some basic knowledge of the physics of STM is necessary to judge the relevance
of experiments. The tunneling junction of an STM is a quantum mechanical
1 SXM: An Introduction 3
system; hence a basic knowledge of quantum mechanics is required to under
stand the physics. An overview of methods and approximations used to model
the tunneling process in STM is given by Baratofi (1984).
1.2.1.1 THE TUNNELING CURRENT-A SIMPLE THEORY
To get a first intuitive view about electron tunneling between the tip and the
sample of an STM, I will consider the textbook case of quantum mechanical
electron tunneling between infinite, parallel, plane metal electrodes. I only treat
the simplest case with no time-dependent potentials. Excellent articles on this
subject have been published by Simmons (1963), (1964), Gundlach (1966),
Brinkman et al (1969), Duke (1969), Hartman (1984), and Teague (1986). The
axis perpendicular to the plane parallel electrodes is the z-axis, with its zero on
the left side of the tunnel gap (see Fig. 1.1).
The electron motion is governed by the Schrodinger equation
/|ψ(ζ,ί) = ΗΨ(ζ,ί), (1.1)
z=0
FIGURE 1.1 Coordinate system for calculating the transmissivity of a one-dimensional tunneling
barrier. The electron plane wave is incident on the barrier from the left (negative z-axis). The two
electrodes are separated by the distance d.
4 Othmar Marti
where Η is the Hamiltonian of the system. The Hamiltonian for a simple tunnel
junction consists of a kinetic energy part
and a potential energy part
ν(ζ)Ψ(ί,ί).
The potential energy is equal to zero everywhere except in the barrier between
the electrodes, from 0 to d, where d is the thickness of the barrier. The wave
function Ψ(ζ, t) of the electrons is a solution of the equation
2
d . Ψ d , . .
i - Ψ(£, t) — — Ψ(ζ, t) + V(z) Ψ(ζ, t)
2
2 A -
( Ψ d
The probability of finding a particle described by the wave function Ψ(ζ, ί)
at the position ζ at the time t is
P(z, t) = Ψ(ζ, 4 Ψ*(ζ, t) = |Ψ(ζ, (1.3)
To simplify the calculation, consider the one-dimensional case of a tunneling
barrier with a potential independent of time. The wave function Ψ(ζ, ή is written
as the product Ψ^(ζ) Ψ,(£). Equation 1.2 can then be separated and written as
0 = |^ Ψ,(ζ) + (£ - V) Ψ,(ζ). (1.4)
We assume that the electrons are incident on the barrier from the left. There
are three solutions to the Schrodinger equation: at the left of the barrier, in the
barrier, and at the right. Our answer to this problem is
'Ae^ + Be-**/*, <0
z
Ψ(ζ) = < Ce-** + De**, 0 < ζ ^ d (1.5)
ζ
I A S ( E ) e * * z - z > d,
where p = V2w£, 1)k = V2m(V — E). (See, for instance, Baym [1969] for a de
tailed treatment of the problem.) At the boundaries of the three regions, these
functions and their first derivative must be continuous. The function S(E) is
called the tunneling matrix element. It is a measure for the probability to tunnel
from left to right for a particle being present at the left side of the junction.
Satisfying the boundary conditions in Eq. 1.5 leads to four simultaneous
equations for the five parameters A, B, C, D, S(E). We can chose an arbitrary
value for the amplitude of the incoming electron wave; hence we set A — 1. The
tunneling matrix element for Ε < V is
