Micron 38 (2007) 824–833

www.elsevier.com/locate/micron

Atomic force microscopy probing of cell elasticity

Tatyana G. Kuznetsova a, Maria N. Starodubtseva a,*, Nicolai I. Yegorenkov b,
Sergey A. Chizhik c, Renat I. Zhdanov d
a
Gomel State Medical University, 5, Lange str., Gomel 246000, Belarus
b
Gomel State Technical University, 48, ave. Oktyabrya, Gomel 246746, Belarus
c
A.V. Lykov Heat and Mass Transfer Institute of NASB, 15, Brouki str., Minsk 220072, Belarus
d
Institute of General Pathology and Pathophysiology, Russian Academy of Medical Sciences, 8, Baltiiskaya St., Moscow 125315, Russian Federation

Abstract
Atomic force microscopy (AFM) has recently provided the great progress in the study of micro- and nanostructures including living cells and
cell organelles. Modern AFM techniques allow solving a number of problems of cell biomechanics due to simultaneous evaluation of the local
mechanical properties and the topography of the living cells at a high spatial resolution and force sensitivity. Particularly, force spectroscopy is used
for mapping mechanical properties of a single cell that provides information on cellular structures including cytoskeleton structure.
This entry is aimed to review the recent AFM applications for the study of dynamics and mechanical properties of intact cells associated with
different cell events such as locomotion, differentiation and aging, physiological activation and electromotility, as well as cell pathology. Local
mechanical characteristics of different cell types including muscle cells, endothelial and epithelial cells, neurons and glial cells, fibroblasts and
osteoblasts, blood cells and sensory cells are analyzed in this paper.
# 2007 Elsevier Ltd. All rights reserved.

Keywords: Atomic force microscopy; Cell elasticity; Young’s modulus; Cytoskeleton dynamics

1. Introduction AFM is a novel method for high-resolution imaging of any

surface including those of living and fixed cells (Bischoff and
It is now well accepted that the cell functions are essentially Hein, 2003, 2004, 2005). This powerful technique is also used for
determined by its structure. At different hierarchy levels the characterization of the mechanical, electrical and magnetic
structural organization of cells is characterized by certain characteristics of samples to be studied both qualitatively and
mechanical properties. It is obvious that cell structure should be quantitatively. AFM operation is based on the detection of
different both in a variety of physiological processes (such as cell repulsive and/or attractive surface forces. The interaction
differentiation, growth, adhesion, etc.) and under pathogenesis between the sample surface and a tip (probe) located very close
(oxidative stress, attack of viruses, parasites, etc.). For a long time to it corresponds to the force between the atoms of the sample and
there have been two approaches for the study of cells’ mechanical those of the tip that scans its surface. Image contrast is generated
properties: (i) the cell mechanical properties were integrally by monitoring the forces of interaction between the tip and the
studied, when the cell was considered as a single whole and (ii) surface. The tip is fabricated under a flexible cantilever
mechanical properties of the cell structural components were responsible for the signal transduction. The interaction between
studied in details using isolated lipid bilayers, biomembrane and the sample and the tip causes bending or twisting of the cantilever
cytosolic proteins. Only recently it has become possible to probe in a manner proportional to the interaction force. A small laser,
micro- and nanomechanical properties of cell structures and to which is focused on the cantilever, detects any bending or
study the spatial distribution of mechanical properties of cellular twisting of the cantilever. The reflection of the laser beam is
structures within a single cell by the atomic force microscopy focused on a photodiode detector. The interaction of the sample
(AFM) technique (Hansma, 2001). with the tip is measured by the variation in the reflected beam’s
point of incidence on the photodiode. Deflection of the cantilever
* Corresponding author at: Gomel State University, 5, Lange str., Gomel
by interaction with features on the sample surface is monitored
246000, Belarus. Tel.: +375 232 745274; fax: +375 232 749831. during scanning and is translated into a three-dimensional image
E-mail address: marysta@mail.ru (M.N. Starodubtseva). of the surface (Mozafari et al., 2005).
0968-4328/$ – see front matter # 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.micron.2007.06.011
 T.G. Kuznetsova et al. / Micron 38 (2007) 824–833 825

AFM can be operated in a number of different imaging usually performs force-curve analysis. The force value versus
modes depending on the nature of the interaction between the distance between the probe and the surface can be plotted in this
tip and sample surface. The micromechanical properties of cell case. The force curve contains information about long- and
surface and subsurface layers can be detected either by contact short-range interactions and represents a basis for estimation of
mode AFM techniques (force modulation, lateral force sample Young’s modulus. Today, there is a serious problem in
microscopy and force-curve analysis) or by phase imaging in estimation of absolute value of cellular Young’s modulus using
the tapping mode AFM (intermittent, semi contact) (Fig. 1). AFM force-curve analysis due to the problem of what
Probing of cell surface by AFM techniques can reveal appropriate mechanical model to choose.
heterogeneities of mechanical properties of the surface at Up to now the Hertz model has been used in the majority of
nanolevel, and subsurface layers of cells. The resolution of articles devoted to the evaluation of Young’s modulus of cells.
AFM in air at vertical direction is 0.1–0.5 nm, and at The Hertz model describes the simple case of elastic
horizontal direction is 1–5 nm, depending on sample rigidity. deformation of two perfectly homogeneous smooth bodies
The horizontal resolution can be solved for living cells in touching under load (Hertz, 1881; Johnson, 1985). Two
aqueous medium even at several tens of nanometers range due important assumptions of the Hertz model are the followings:
to the softness of the cell membrane. The thickness of cellular (i) the indenter must have a parabolic shape and (ii) the indented
membranes is known to be around 5–10 nm. When analyzing sample is assumed to be extremely thick in comparison to the
their heterogeneities by AFM techniques we are able to create indentation depth. The first assumption remains a valid one for
the picture of the cellular structure of certain regions within a the case when a spherical tip radius is much bigger than the
single cell. The sensitivity and resolution of AFM method indentation depth (h < 0.3R) (Mahaffy et al., 2000).
depend also on tip and cantilever characteristics (e.g., radius, If the tip of an AFM nanoscope is approximated by a sphere
shape, material) (Alessandrini and Facci, 2005). with the radius R, then the force on cantilever F(h) is given by:
The basic AFM technique for quantitative study of pffiffiffi
mechanical characteristics of cells and tissues is the force 4 R  3=2
FðhÞ ¼ E h
spectroscopy (namely, force-curve analysis). By recording the 3
force value and vertical deflection of the cantilever, the probe where h is the depth of the indentation, E* the effective
approaches the surface under the study at the fixed point and modulus of a system tip-sample, which is calculated from

Fig. 1. Scheme of atomic force microscopy (AFM) probing the cells.

826 T.G. Kuznetsova et al. / Micron 38 (2007) 824–833

the equation: the Hertz model reflects the same tendency, but has different
means (0.87  0.23 and 1.75  0.43 kPa for control and
1 1  v2tip 1  v2sample sheared endothelial cells, respectively).
¼ þ
E Etip Esample The AFM experimental approach named as force integration
to equal limits (FIEL) mapping, for producing quantitative
in which Etip, vtip and Esample, vsample are the Young’s modules
maps of relative cell elasticity was developed in 1998 (A-
and the Poisson ratios for the materials of tip and the sample,
Hassan et al., 1998). FIEL theory assumes a simple relationship
respectively. If the material of the tip is considerably harder
between values of the works done by the AFM cantilever during
than the sample the following equation is used (Vinckier and
an indentation and the elastic constants at different surface
Semenza, 1998):
positions.
Esample The collection of force curves over a certain area allows
E 
1  v2sample creation of the elasticity map of the cell surface. The surface
elasticity map can be also obtained using either force-
The Sneddon’s variation of the Hertz model is used for the modulation (static mode) or phase imaging (tapping mode)
case of cone tip of AFM cantilever (Laurent et al., 2005): techniques. The characteristics of cantilever oscillation
2 Esample (amplitude and phase shift) carry on information about local
FðhÞ ¼ tan a h2 elastic and friction properties of the sample in both cases. The
p 1  v2sample
image of the changes in oscillation characteristics represents a
where a is the half-opening angle of the AFM tip. map of relative mechanical properties of cell surface.
Though the indentation depth in case of AFM probing of the Indirect information on elastic properties of cell surface can
cell is in the range of hundreds of nanometers, which is higher be provided by recording the lateral force map of surface to be
than an appropriate depth for the Hertz model, it was shown in studied. The AFM cantilever lateral deflections (torsion) arise
many studies that the Hertz model describes sufficiently the because of either the changes in the surface slop or heterogeneity
experimental data. The original Hertz theory did not allow of surface frictional properties. The lateral force map is
adhesion of the indenter to material. Johnson, Kendal and simultaneously analyzed with sample surface topography to
Roberts modified the theory for that case (Johnson et al., 1971). elaborate the specificity of elastic property map.
The Hertz model was mainly used for an estimation of static This paper is aimed to analyze the progress in the usage of
Young’s modulus of cells, but dynamic Young’s modulus was AFM probing elasticity of mammalian cells for the study of their
sometimes used for the characterization of cell elastic temporal and spatial structural dynamics under physiological and
properties (Mahaffy et al., 2004). Because cell surface is pathological processes. To analyze the mechanical properties of
heterogeneous (that is a network of cell membrane and sub- different cells comparatively, we only chose the articles in which
membrane structures), the cellular Young’s modulus evaluation Young’s modulus were used for the characterization of cell
using the Hertz model assumes an error. mechanical properties.
The second model used for studying cell elastic properties
basing on force spectroscopy data is a model based on the 2. Young’s modulus and its AFM measurements in
theory of elastic shells (ES) (A-Hassan et al., 1998; Scheffer living cells
et al., 2001; Timoshenko and Woinowsky-Krieger, 1970). This
theory considers cells as the shells filled with liquid. In such an Table 1 summarizes the results of Young’s modulus
approach, effective Young’s modulus can be evaluated from the measurement for different types of living mammalian cells.
relationship between effective Young’s modulus, shell thick- It shows that the elastic modulus value of living cells varies in
ness and bending modulus. The serious problem of such wide range. It is evident that both real variability of the
evaluation procedure is the determination of the boundary parameter and imperfection of AFM methods of measurement
conditions for the calculation of any constants involved in main and numerical estimation of cell elasticity are present. The
relation and the definition of the tip-sample contact radius. analysis of almost a decade progress in this area allows
Among other theoretical models used in AFM-based generalizing some methodological factors having a significant
evaluation of cell mechanical characteristics we should influence on Young’s modulus value. The technical and
mention the finite element model, which is the most popular theoretical problems of those studies were discussed in the
model for analysis of elasticity problems in engineering Section 1. Here, we discuss the problems connected with cell
(Ohashi et al., 2002). specificity. Literature survey demonstrated that under a change
The values of elasticity parameters calculated using various of the external conditions, the elasticity of cell membranes
models differ each other. For example, Ohashi’s group studying changes much stronger than the morphology of cell.
mechanical properties of endothelial cells exposed to shear The first factor is a question of AFM sample handling. The
stress (Ohashi et al., 2002) calculated different values of appropriate object for an illustration of the progress in this acute
Young’s modulus using various models. The modulus values area is erythrocytes. AFM method has been used to study both the
calculated using a finite element model appeared to be living (Nowakowski et al., 2001; Kamruzzahan et al., 2004) and
significantly higher: from 12.2  4.2 to 18.7  5.7 kPa with fixed erythrocytes either in air (Gould et al., 1990) or in buffer
exposure to shear stress. The modulus value calculated using (Butt et al., 1990). The living cells are rather soft and delicate for
 T.G. Kuznetsova et al. / Micron 38 (2007) 824–833 827

Table 1
Young’s modulus of mammalian cells
Cell type E (kPa) Commentary References
Endothelial cells
HUVEC 10–11 Sato et al. (2004)
– 1.3–7.2 Spatial heterogeneity Mathur et al. (2000)
– 0.9–1.7; 12.0–18 Shear stress, different models Ohashi et al. (2002)
BPAEC 0.2–2.0 Spatial heterogeneity Pesen and Hoh (2005)
Leukocytes
Leukemia myeloid cells (HL60) 0.2–1.4 Rosenbluth et al. (2006)
Leukemia lymphoid (Jurkat) cells 0.02–0.08
Neutrophils 0.2–0.07
Corti organ’s cells
Outer hair cells 300–400 Cortical lattice Tolomeo et al. (1996)
Guinea pig’s outer hair cells 2–4 Different levels of cochlea Sugawara et al. (2002)
Mouse outer hair cells 2–4 Murakoshi et al. (2006)
Guinea pig’s inner hair cells 0.1–0.5 Sugawara et al. (2002)
Hensen’s cells 0.3–1.1 –
Osteoblasts 0.3–20.0 Changes at adhesion Simon et al. (2003)
Astrocytes 2–20 Spatial heterogeneity Yamane et al. (2000)
Fibroblasts 4–5 Bushell et al. (1999)
Migrating 3T3 cells 3–12 Spatial heterogeneity Rotsch et al. (1999)
– 0.6–1.6 Changes at adhesion Mahaffy et al. (2004)
L 929 4–5 Wu et al. (1998)
Epidermal keratocytes 10–55 Spatial heterogeneity Laurent et al. (2005)
Platelets 1–50 Spatial heterogeneity at activation Radmacher et al. (1996)
Skeletal muscle cells
Murine C2C12 myoblasts 11–45 Change at differentiation Collinsworth et al. (2002)
Murine C2C12 myotubes 8–14 Changes at differentiation, Treating with L-arginine Zhang et al. (2004)
– 10–17 –
– 28–21 Mathur et al. (2001)
Myofibrils 40–45 Yoshikawa et al. (1999)
Cardiocytes 90–110 Mathur et al. (2001)
Rat 32–42 Aging changes Lieber et al. (2004)
Chicken 5–200 Spatial heterogeneity Hofmann et al. (1997)
Erythrocytes 14–18 Mozhanova et al. (2003)
19–33 Normal Dulinska et al. (2006)
22–64 Hereditary spherocytosis, thalassemia, G6PD deficiency –
16–64 –
70–110 –

their AFM probing under physiological conditions. Their drying, solutions leads to smearing of the AFM image and the maximal
freezing and fixing with chemical agents improve the AFM space resolution of living erythrocytes can be only about 200 nm
images and AFM indentation results. However, these procedures (Mozhanova et al., 2003).
change cell structure, viability and elasticity. The Young’s Standard AFM technique for cell elasticity measurement is
modulus values for erythrocytes treated with 5% formalin based on indentation of the cells firmly attached to the
solution are increased 10-fold (119.5  15 kPa) compare to substrate. For reliable results of indentation, the firm substrate/
viable (native) erythrocytes (16.05  2.3 kPa) (Mozhanova cell contact is required and that is a problem for nonadhered
et al., 2003). Transverse stiffness of cardiomyocytes is also cells in solution. A good approach for the immobilization of
increased by a factor of 16 after fixing with formalin (Shroff et al., native erythrocytes in liquid is attachment to glass surface
1995). Takeuchi et al. (1998) comparing a variety of methods for previously modified with poly-L-lysine solution. Poly-L-lysine
preparing erythrocyte ghost for AFM studies showed that air provides the accurate localization of red blood cells on the glass
drying is not suitable even after fixation in glutaraldehyde. On the surface due to electrostatic interaction between the negatively
other hand, fixation enhances the images of cell structures like charged cell surface and the positively charged poly-L-lysine
the cytoskeleton (Shroff et al., 1995, Hofmann et al., 1997). layer. However, poly-L-lysine can induce membrane rearrange-
Moreover, the highest resolution for cells, 10 nm, may be ment with the formation of specific membrane deformation
achieved only in air, which presumes cell fixation before AFM pattern within contact area (Dulinska et al., 2006). Recently, a
probing. The high mobility of the erythrocyte shape in buffer new method of indentation of leukemia cells placed at special
828 T.G. Kuznetsova et al. / Micron 38 (2007) 824–833

microwells was reported (Rosenbluth et al., 2006). This method found in mechanically active environment, and they are
provides the mechanical immobilization of cells, but also has an required to withstand shear stress, blood pressure, and any
influence on the estimated Young’s modulus value. In this case changes in pressure due to breathing cycles. The earliest AFM
the cell deformation is well described by elastic model based on works on living aortic endothelial cells were devoted to
Hertzian mechanics. studying the effects of shear stress on cellular organization and
The second factor is related to the heterogeneity of other factors which may influence the mechanical response of
mechanical properties of cells. There are significant variations cells to flow (Barbee et al., 1994; Barbee, 1995; Sato et al.,
of the values of elastic modulus at different cell regions. Mathur 2000; Ohashi et al., 2002). Ohashi et al. (2002) reported that the
et al. (2000) showed that the elastic modulus value of human local elastic parameters of aortic endothelial cells significantly
umbilical vein endothelial cells was 7.22  0.46 kPa over the increased (from 0.87  0.23 to 1.75  0.43 kPa) with exposure
nucleus; 2.97  0.79 kPa over the cell body in proximity to the to shear stress. The average elastic modulus values of bovine
nucleus, and 1.27  0.36 kPa on cell body near the edge. Costa pulmonary artery endothelial cells (BPAECs) reported by Pesen
and Yin (1999) reported that the cell body of bovine pulmonary and Hoh (2005) were in the similar range of 0.2–2 kPa.
artery endothelial cells was two- to three-fold softer than the Miyazaki and Hayashi (1999) demonstrated the difference in
cell periphery. The corresponding study on cardiomyocytes the mechanical properties of rabbit endothelium. So, cells were
also revealed that cells are softer at the nuclear region, and stiffer in the medial wall of aortic bifurcation than in the lateral
become stiffer toward the periphery (Shroff et al., 1995). wall.
Mapping of the Young’s modulus across the living chicken Using AFM and human umbilical vein endothelial cells
cardiocytes, Hofmann et al. (1997) stated that the stress fibers (HUVEC), Mathur et al. (2000) showed that the cell responds
were characterized by the presence of areas with a stiffness of globally to the localized applied force over the cell edge and the
100–200 kPa embedded in softer parts of the cell, with elastic nucleus. They concluded that the nuclear region of the cell
modulus values between 5 and 30 kPa. The elasticity map appears to be stiffer than the rest of the cell body and although
images of living astrocytes (glial cells of nerve tissue) showed the nucleus appears to be offset from the basal surface, the focal
that cell membrane above the nucleus was softer (2–3 kPa) than adhesion movement upon the apical cell surface perturbation
the surroundings, and that the cell membrane above ridge-like indicates a link between the nucleus and the focal adhesions via
structures reflecting F-actin was stiffer (10–20 kPa) than the the cytoskeleton (Mathur et al., 2000).
surroundings. In the elasticity map images of fixed astrocytes,
on the other hand, the elasticity value of cells was found to be 3.2. Platelet activation
relatively uniform (200–700 kPa) irrespective of the inner
structures of cells (Yamane et al., 2000). On the other hand, the Activation of platelets, leading to a drastic change in the
variations of elastic modulus value within single cell are often cytoskeletal structure and marked change in cell shape, can be
objects of the study themselves. induced by a contact with wettable surfaces or with tip during
The third factor influencing elastic modulus measurement is AFM probing. Fritz et al. (1994, 1993) demonstrated that
connected with cell thickness. Substrate contributions can be platelets, which have bound to the surface but not yet activated,
neglected if AFM tip never indented more than 10% of the cell can be scanned by AFM at low forces without any noticeable
thickness (Mathur et al., 2001). If the cell compartment under the time-dependent changes in shape. However, scanning at higher
study is very thin (<1000 nm), e.g., in the case of lamellipodium, forces promotes their activation. Topographic and elasticity
it is necessary to impart special challenges for accurate maps of viable-activated cells were created and analyzed using
measurements of its viscoelastic behavior. Mahaffy et al. force mapping techniques (Radmacher et al., 1996). The
(2004) reported the method for AFM-based microrheology that authors reported that the elastic modulus values of activated
allowed to estimate the viscoelastic constants of thin parts of cell platelets were in a range of 1–50 kPa.
(<1000 nm) as well as those of thick areas, applying two
different models—a model for well-adhered regions and a model 3.3. Cell locomotion
for nonadhered regions.
Cell motility is ultimately a mechanical phenomenon, but
3. Cell biology and cell elasticity very little is known about the mechanical and physical
properties of moving cells, including these properties in
Although Young’s modulus obtained by AFM techniques specific cell regions at specific stages of the cell migration
must be carefully assessed as the absolute values, it is very useful process. So, nanoindentation can be very useful in this case.
as relative parameter in certain experiments. Therefore, Young’s A few AFM studies of time-dependent structural changes, cell
modulus can be successfully used in the study of a variety of cell migration and the associated changes in mechanical properties
functions, some examples of which are given below. have been reported for fibroblasts (Sasaki et al., 1998; Nagayama
et al., 2001; Haga et al., 2000). The usage of the force modulation
3.1. Functional mechanics of endothelial cells mode allowed demonstrating the existence of a correlation
between the time-dependent changes of cell surface and the
Vascular endothelial cells represent an interesting system for elastic parameters of viable mouse fibroblasts in culture medium
studying cell mechanics and cytoskeleton itself. These cells are (Sasaki et al., 1998). It is interesting that some cells continued to
 T.G. Kuznetsova et al. / Micron 38 (2007) 824–833 829

shrink and change their softness for several hours. The results during adhesion (Domke et al., 2000; Takai et al., 2005, Simon
indicated that the AFM force-mapping technique does not et al., 2003). Liang et al. (2004) employed AFM and model
appreciably perturb cell mechanical dynamics. cells for the elucidation of micromechanical properties of
A clear relationship between the local stiffness distribution biomembranes (Fig. 2). Domke et al. (2000) used the AFM
on cell and cell migration was also found. The stiffness method for testing the biocompatibility of implanted materials
distribution of cell surface was quite constant for stationary by the study of the elastic properties of osteoblast cells adhered
cells, but if cells started to move, the stiffness in their nuclear to the different surfaces. Takai et al. (2005) determined, e.g., the
regions was drastically decreased (Nagayama et al., 2001). apparent elastic modulus of osteoblast-like MC3T3-E1 cells
The mechanical properties of the leading edge of migrating adhered to different surfaces. They stated that the elastic
cells, i.e., the lamellipodium, are important for a deeper modulus values of osteoblasts adhered to extracellular matrix
understanding of cell locomotion mechanism. Haga et al. proteins that bind cells using integrins were higher compared to
(2000) found that the stable part of the fibroblast cell body was cells on glass and poly-L-lysine adhering the cells through
stiffer and did not demonstrate morphological changes for over nonspecific binding. It was suggested that the enhanced
1 h. The authors proposed that it is connected with excess modulus value of osteoblasts adhered to extracellular matrix
condensation of the actin network, hardening cell cortex, and proteins was due to remodeling of the actin cytoskeleton, and
lowering cytoskeletal activity. The nuclear region of cell body modulation of cell stiffness upon adhesion to various substrates
was slightly less viscous than the peripheral region. may influence mechanosignal transduction in osteoblasts. The
Dimensional and mechanical dynamics of active and stable elastic properties of osteoblasts cultured on two types of surface
edges in motile 3T3 fibroblasts in culture were also investigated to induce weak and strong cellular adhesions were also studied
(Rotsch et al., 1999). The cortical stiffness calculated for the by AFM technique (Simon et al., 2003). The values of elastic
stable edges was 12 kPa. Contrary to that, the leading edge modulus were between 0.3 and 200 kPa depending on the level
stiffness had an upper limit of 3–5 kPa. of cell adhesion and the approaches used to measure this elastic
Recently, the fish epidermal keratocytes with their stable modulus value. The authors concluded that a comparison
shape and steady motion were studied (Laurent et al., 2005). between the elasticity of viable cells may be used as
The authors demonstrated that though vertical lamellipodia demonstration of cytoskeletal reorganization and the state of
thickness remained nearly constant, the rigidity value was cell adhesion.
highest at the front protruding edge of the lamellipodia, and Studying the elastic properties of fibroblast lamellipodium,
gradually decreased along with a distance from the edge. The Mahaffy et al. (2004) applied two different models: a model for
values of the elastic modulus exhibited a tendency to decrease well-adhered regions and a model for nonadhered regions. They
from 55 kPa at the front of the lamellipodium to 10 kPa at its showed that very thin regions relatively near the edge of NIH
rear. Those differences in rigidity may indeed reflect the 3T3 fibroblasts were strongly adherent with an elastic strength
differences in the structure of the leading edge between value of 1.6  0.2 kPa, and the regions quite far from the edge
different types of fibroblasts. of these cells adhered worse, and therefore Young’s modulus
value was less (0.6  0.1 kPa).
3.4. Cell adhesion
3.5. Physiology of sensory cells
AFM method is also effective for the study of cell
mechanical properties during cell adhesion. There are some The mechanical property measurement can also be useful for
AFM studies which characterized osteoblast elastic properties the study of sensory cells physiology including sensory hearing

Fig. 2. A representative AFM height image of fixed model cell membranes (EggPC:DCP:Chol, 7:2:1 molar ratio). Imaging was done at a scan rate of 0.3 Hz.
Courtesy of Dr. M.R. Mozafari.
830 T.G. Kuznetsova et al. / Micron 38 (2007) 824–833

cells, capable to alter their length in response to changes in of cell functions thoroughly. To provide the fast progress in
membrane potential and subject the basilar membrane of inner studying cellular mechanisms with cytoskeleton involvement,
ear to force, resulting in cochlear amplification. The force AFM analysis is often combined with confocal fluorescent
produced by piliform (hear hair) cells’ electromotility is microscopy or usage of a drug destructing cytoskeleton
thought to depend not only on the conformational change of the components. The data collected show clearly that elastic
protein motors, but also on the mechanical properties of the cell response of cell is due to the actin network to a high degree.
lateral wall (Murakoshi et al., 2006). Sugawara et al. (2002) Using all these methods Pesen and Hoh (2005) characterized the
revealed that elastic modulus values in the apical region of the micromechanical architecture of the cortex in bovine pulmonary
outer piliform cells were three times higher than those in the artery endothelial cells. The authors showed that the cortex in
basal and middle regions. Other reports (Wada et al., 2003) also these cells is organized as a polygonal mesh at two levels: a
state that the stiffness of the apical region of outer piliform cells coarse mesh with dimensions of several micrometers and a fine
is greater than that in other regions. According to Wada’s overlapping mesh with dimensions of hundreds of nanometers.
results, a difference between the intervals of the actin These meshes appear to be intertwined and are in part composed
circumferential filaments in the apical region and those in of actin and vimentin. The analysis of fluorescent images and
other regions is one factor that causes the high stiffness in this elasticity maps revealed that in the case of activated platelets
part of cell lateral wall. It was found that Young’s modulus (Radmacher et al., 1996) and cardiomyocytes (Shroff et al.,
value decreases with an increase in the piliform cell length. 1995) the variation of elastic properties across the cell was
Young’s modulus values in the middle region of a long outer correlated with the cytoskeletal heterogeneity as well as the
piliform cells obtained from the apical turn of the cochlea and significant increase in the elastic modulus value of endothelial
that of a short outer piliform cells obtained from the basal turn cells with exposure to shear stress occurred due to the remodeling
or the second turn were 2.0  0.81 kPa and 3.7  0.96 kPa, of cytoskeletal structure (Ohashi et al., 2002).
respectively (Sugawara et al., 2002). The role of cytoskeletal elements in formation of the
The calculated Young’s modulus values of the guinea pig cells mechanical properties of cells was found more precisely using a
were 0.29  0.20 kPa for inner hair cells and 0.69  0.45 kPa for variety of medicines affecting the cytoskeleton. According to
Hensen’s cells (Sugawara et al., 2004). It is interesting that the Rotsch et al. (1997), who studied viable cultured rat liver
species differences in the estimated values of elastic modulus of macrophages, the chemical disassembly of the actin network by
each type of hair cells is insignificant. So, the Young’s modulus applying cytochalasin B decreases the cell’s average elastic
value of the mouse outer hair cells in the apical turn of the cochlea modulus seven-fold within less than 40 min. Treating cells with
(2.1  0.5 kPa) was similar to that of the guinea pig cochlea latrunculin A results in a two-fold decrease in the elastic
(Murakoshi et al., 2006). modulus of the perinuclear region after 40 min, whereas other
parts of cell are not affected. The latruculin-induced disruption
3.6. Role of cytoskeleton in elastic property formation of the cytoskeleton network was also observed by AFM images
of skin fibroblasts (Braet et al., 2001) and two different
The unique advantage of AFM method is the opportunity to fibroblast cell lines (Rotsch and Radmacher, 2000). A decrease
perform measurement of the mechanical properties of cells and of the elastic modulus for viable chicken cardiocytes after their
visualization of important cellular structures like cytoskeleton treatment with cytochalasin B as well as for L929 cells affected
simultaneously (Fig. 3). It allows understanding the mechanisms with cytochalasin D was demonstrated by two groups

Fig. 3. The unique structure of lamellipodium cytoskeleton determines its local mechanical properties: (a) topography of small lymphocyte (5 mm  5 mm) and (b)
lateral force map of its lamellipodia (1.5 mm  1.5 mm). Arrows show the structures arranged transverse to lamellipodium growth direction. The lymphocytes were
isolated from peripheral venous blood obtained from healthy volunteers by centrifugation over a Ficoll gradient. The cells were treated with 0.5 mM peroxynitrite in
suspension, placed on glass plate for 20 min adhesion, fixed with 1% glutaraldehyde, dried on an air. The topography and lateral force map of lymphocyte surface
were produced by AFM ‘‘NT-206’’ (Belarus).
 T.G. Kuznetsova et al. / Micron 38 (2007) 824–833 831

(Hofmann et al., 1997; Wu et al., 1998). On the contrary, the the time course of differentiation of myoblast to myofiber. They
treatment of fibroblasts with nocodazole or colcemid induces a demonstrated the dramatic change in the passive mechanical
marked increase in their elasticity (Wu et al., 1998). Using such behavior of mouse skeletal myocytes which appeared in a
cytoskeleton medicines as cytochalasins B and D, latrunculin A significant increase of the apparent elastic modulus with
and Jasplakinolide, Rotsch and Radmacher (2000) showed that differentiation from 11.5  1.3 kPa for undifferentiated
disaggregation of actin filaments results in a decrease in the myoblasts to 45.3  4.0 kPa after 8 days of differentiation.
elastic modulus of fibroblasts, while reorganization of Results of examining the actin, myosin, and tubulin
microtubules did not affect cell elasticity. demonstrated that major contributors to changes in the transverse
The rigidity profile of migrating epidermal keratocytes elastic modulus during differentiation are actin and myosin.
closely resembles the actin density profile, suggesting that the The elastic modulus of the multinucleated myocytes and
dynamics of rigidity is due to actin depolymerization (Laurent its varieties during cell differentiation were also estimated
et al., 2005). The authors suggest that a decrease of rigidity may (Zhang et al., 2004). As it turned out at the second day of cell
play a role in facilitating the contraction of the actin-myosin differentiation, elastic modulus for statically stretched cells
network at the lamellipodium/cell body transition zone. Haga was 8.3  1.6 kPa and increased until fourth day up to
et al. (2000) also propose that higher rigidity of cell body 14.3  2.4 kPa.
compared to lamellipodia of migrating fibroblast is connected
to an excess condensation of the actin network, hardening the 3.8. Pathology
cell cortex, and lowering the cytoskeletal activity.
AFM investigations can be useful for studying cell
3.7. Cell differentiation and aging pathology. Any factors having an influence on cell structures
can cause the alterations in mechanical properties of cell
The changes in mechanical properties and cytoskeleton (Garcia et al., 1997). The determination of the local elastic
reorganization seem to be correlated with cell cycle stages properties of cells in their culture conditions has opened the
(Sato et al., 2004; Collinsworth et al., 2002; Zhang et al., 2004; possibility for the measurement of the influence of different
Lieber et al., 2004; Berdyyeva et al., 2005). Therefore, these factors on the mechanical properties of the living cells. That is
results form the basis for understanding the mechanisms of cell why the AFM estimation of cell mechanical properties seems to
differentiation and organism ageing. be a perspective method of diagnostics of different pathologies.
AFM studies of human umbilical vein endothelial cells Dulinska et al. (2006) studied the elastic properties of
reveal that cell elasticity depends on the culture period (Sato erythrocytes from patients with different types of anemias using
et al., 2004). The elasticity of cells cultured on type IV collagen force spectroscopy and compared the results with those obtained
for longer than 4 days leads to average elasticity values higher in normal cells. Additional comparison was performed for
than 10 kPa. The data obtained for human epithelial cells anisocytic erythrocytes since the authors considered that
(Berdyyeva et al., 2005) also prove an increase in cell rigidity alteration of the erythrocytes shape could be reflected by
during ageing in vitro. changes of Young’s modulus. According to Dulinska et al. (2006)
AFM-nanoindentation has been recently used to analyze the Young’s modulus of pathological erythrocytes was two to three
ageing changes of cardiac myocytes of young and old male times higher than in normal cells. The values of elastic modulus
Fischer 344  Brown Norway F1 hybrid rats (Lieber et al., were: 26  7, 43  21, 40  24 and 90  20 kPa for control,
2004). The significant increase of the apparent elastic modulus hereditary spherocytosis, thalassemia, and G6PD deficiency,
of cardiac myocytes with advanced age was found. The elastic respectively. The maximal change was observed for erythrocytes
modulus values are changed from 35.1  0.7 kPa for young rat with G6PD deficiency, where the calculated Young’s modulus
cells to 42.5  1.0 kPa for old rat cells. Results of the study was more than three times larger than in normal cells. The authors
support the authors’ concept that the mechanism mediating LV attribute the increase of the Young’s modulus to the change in
diastolic dysfunction in ageing hearts resides, in part, at the cytoskeleton structure whether due to the changes of spectrin
level of the myocyte. structure, the molecular anomaly in hemoglobin structure or
The differentiation of skeletal muscle is a complex process, impaired ATP metabolism. They also showed the change in
which helps to understand the functional properties and distribution of Young’s modulus in erythrocytes. It became
mechanisms of the muscle tissue regeneration. It includes broader in case of pathologically altered erythrocytes and even
subsequent fusion of myoblasts to form multinucleated became bimodal at anisocytosis.
myotubes or myofibers, and expression of differentiation- In our laboratory the comparative AFM study of elastic
specific proteins. Cell transformation is accomplished by the properties of normal and peroxynitrite-treated erythrocyte was
changes of cytoskeletal structures and probably lead to the carried out. Peroxynitrite as a reactive nitrogen species reacts
changes in the mechanical properties of differentiated and with both protein and lipid components of membrane and
undifferentiated skeletal muscle cells. membrane skeleton. According to our data peroxynitrite
Collinsworth et al. (2002) used AFM to elucidate the nature of (>1 mM) causes about two- to three-fold increase in Young’s
mammalian myocyte mechanical properties throughout their modulus of erythrocyte and also results in the broadening of the
development and to test the hypothesis that the transverse elastic Young’s modulus distribution peak. We also attribute these
and viscous properties of skeletal muscle cells change throughout changes of erythrocyte mechanical properties distribution to
832 T.G. Kuznetsova et al. / Micron 38 (2007) 824–833

cytoskeletal rearrangements. We visualized those cytoskeleton Barbee, K.A., Davies, P.F., Lal, R., 1994. Shear stress-induced reorganisation of
the surface topography of living endothelial cells imaged by atomic force
rearrangements with lateral force map of erythrocyte surface.
miroscopy. Circ. Res. 74, 163–171.
Influence of hydrocortisone on the mechanical and Berdyyeva, T.K., Woodworth, C.D., Sokolov, I., 2005. Human epithelial cells
morphological properties of confluent cell layers of brain increase their rigidity with ageing in vitro: direct measurements. Phys. Med.
microvascular endothelial cells was examined by Schrot et al. Biol. 50 (1), 81–92.
(2005). They showed that hydrocortisone induced a reduction Bischoff, G., Hein, H.-J. (Eds.), 2003. Micro- and Nanostructures of Biological
of the intercellular contact surface and changes in cell Systems, vol. 1. Shaker Verlag, Aachen.
Bischoff, G., Hein, H.-J. (Eds.), 2003. Micro- and Nanostructures of Biological
elasticity. Systems, vol. 2. Shaker Verlag, Aachen.
Because epithelial cells most often are subjected to Bischoff, G., Hein, H.-J. (Eds.), 2003. Micro- and Nanostructures of Biological
carcinogenic transformation, comparative studies of normal Systems, vol. 3. Shaker Verlag, Aachen.
and cancerous human epithelial cells were also carried out Braet, F., de Zanger, R., Seynaeve, C., Baekeland, M., Wisse, E., 2001. A
comparative atomic force microscopy study on living skin fibroblasts
(Lekka et al., 1999). According to this work the normal cells
and liver endothelial cells. J. Electron Microsc. (Tokyo) 50 (4),
had a Young’s modulus of about one order of magnitude higher 283–290.
than cancerous ones. The authors suggested that such change in Bushell, G.R., Cahill, C., Clarke, F.M., Gibson, C.T., Myhra, S., Watson, G.S.,
elastic properties might be attributed to a difference in the 1999. Imaging and force–distance analysis of human fibroblasts in vitro by
organization of cell cytoskeletons. This change is associated atomic force microscopy. Cytometry 36 (3), 254–264.
with the increased crosslinking of extracellular matrix proteins. Butt, H.-J., Wolff, E.K., Gould, S.A.C., Northern, B.D., Peterson, C.M.,
Hansma, P.K., 1990. Imaging cells with the atomic force microscope. J.
In a later work the same authors (Lekka et al., 2001) showed the Struct. Biol. 105, 54–61.
strong correlation between the decrease of the energy Collinsworth, A.M., Zhang, S., Kraus, W.E., Truskey, G.A., 2002. Apparent
production and the increase in Young’s modulus values elastic modulus and hysteresis of skeletal muscle cells throughout differ-
obtained for the cancer cells treated with chitosan. entiation. Am. J. Physiol. Cell Physiol. 283, C1219–C1227.
Costa, K.D., Yin, F.C.P., 1999. Analysis of indentation: implications for
measuring mechanical properties with atomic force microscopy. J. Bio-
4. Conclusions mech. Eng. 121, 462–471.
Domke, J., Dannohl, S., Parak, W.J., Muller, O., Aicher, W.K., Radmacher, M.,
AFM probing of whole cells is an effective tool for studying 2000. Substrate dependent differences in morphology and elasticity of
membrane and sub-membrane cell structures. Because the living osteoblasts investigated by atomic force microscopy. Colloids Surf.
B: Biointerfaces 19 (4), 367–379.
heterogeneity of cell mechanical properties is mainly defined
Dulinska, I., Targosz, M., Strojny, W., Lekka, M., Czuba, P., Balwierz, W.,
by membrane cytoskeleton, AFM probing of the cell elasticity Szymonski, M., 2006. Stiffness of normal and pathological erythrocytes
can be effectively used in investigation of cytoskeleton studied by means of atomic force microscopy. J. Biochem. Biophys. Meth.
characteristics and dynamics, for example, for studying 66, 1–11.
migrating cells. Although for these purposes the force Fritz, M., Radmacher, M., Gaub, H.E., 1994. Granula motion and membrane
spectroscopy methods are commonly used, the lateral force spreading during activation of human platelets imaged by atomic force
microscopy. Biophys. J. 66 (5), 1328–1334.
microscopy, phase imaging and force modulation methods can Fritz, M., Radmacher, M., Gaub, H.E., 1993. In vitro activation of human
be useful in the evaluation of spatial distribution of mechanical platelets triggered and probed by atomic force microscopy. Exp. Cell Res.
and structural characteristics of cytoskeleton. Another per- 205, 187–190.
spective opportunity of AFM probing cell elasticity is Garcia, C.R.S., Takeuschi, M., Yoshioka, K., Miyamoto, H., 1997. Imaging
investigation of the lipid phase characteristics and dynamics Plasmodium falciparum-infected ghosts and parasite by atomic force
microscopy. J. Struct. Biol. 119, 92–98.
on living and fixed cells using phase imaging in intermittent Gould, S.A.C., Drake, B., Prater, C.B., Weisenhorn, A.L., Manne, S., Hansma,
mode. Estimation of structural state (cytoskeleton or membrane H.G., Hansma, P.K., Massie, J., Longmire, M., Elings, V., Northern, B.D.,
lipid phase) of living cells with AFM can be of great importance MuKergee, B., Peterson, C.M., Stoeckenius, W., Albrecht, T.R., Quate, C.F.,
in diagnostics of different human diseases. 1990. From atoms to integrated circuit chips, blood cells, and bacteria with
In this article, we were focused at AFM applications to the atomic force microscope. J. Vac. Sci. Technol. A8, 369–373.
Haga, H., Nagayama, M., Kawabata, K., Ito, E., Ushiki, T., Sambongi, T., 2000.
mainly characterize the mechanical properties of intact cells Time-lapse viscoelastic imaging of living fibroblasts using force modulation
both quantitatively and qualitatively. We realize that only a mode in AFM. J. Electron Microsc. (Tokyo) 49 (3), 473–481.
part of the work done in the field could be considered in a Hansma, H.G., 2001. Surface biology of DNA by atomic force microscopy.
single article, other contributions will be covered in future Annu. Rev. Phys. Chem. 52, 71–92.
papers. Hertz, H., 1881. Ueber den kontakt elastischer koerper. J. fuer die Reine
Angewandte Mathematik 92, 156.
Hofmann, U.G., Rotsch, C., Parak, W.J., Radmacher, M., 1997. Investigating
References the cytoskeleton of chicken cardiocytes with the atomic force microscope. J.
Struct. Biol. 119, 84–91.
A-Hassan, E., Heinz, W.F., Antonik, M.D., D’Costa, N.P., Nageswaran, S., Johnson, K.L., 1985. Contact Mechanics. Cambridge University Press,
Schoenenberger, C.-A., Hoh, J.N., 1998. Relative microelastic mapping of Cambridge.
living cells by atomic force microscopy. Biophys. J. 74, 1564–1578. Johnson, K.L., Kendall, K., Roberts, A.D., 1971. Surface energy and the contact
Alessandrini, A., Facci, P., 2005. AFM: a versatile tool in biophysics. Meas. Sci. of elastic solids. Proc. R. Soc. Lond. Ser. A 324, 301–321.
Technol. 16, R65–R92. Kamruzzahan, A.S., Kienberger, F., Stron, C.M., Berg, J., Huss, R., Ebner, A.,
Barbee, K.A., 1995. Changes in surface topography in endothelial monolayers Zhu, R., Rankl, C., Gruber, H.J., Hinterdorfer, P., 2004. Imaging morpho-
with time at confluence: influence on subcellular shear stress distribution logical details and pathological differences of red blood cells using tapping-
due to flow. Biochem. Cell Biol. 73, 501–505. mode AFM. Biol. Chem. 385 (10), 955–960.
 T.G. Kuznetsova et al. / Micron 38 (2007) 824–833 833

Laurent, V.M., Kasas, S., Yersin, A., Schäffer, T.E., Catsicas, S., Dietler, G., Rotsch, C., Jacobson, K., Radmacher, R., 1999. Dimensional and mechanical
Verkhovsky, A.B., Jean-Jacques Meister, J.-J., 2005. Gradient of rigidity in dynamics of active and stable edges in motile fibroblasts investigated by
the lamellipodia of migrating cells revealed by atomic force microscopy. using atomic force microscopy. Proc. Natl. Acad. Sci. U.S.A. 96, 921–926.
Biophys. J. 89, 667–675. Rotsch, C., Radmacher, R., 2000. Drug-induced changes of cytoskeletal
Lekka, M., Laidler, P., Gil, D., Lekki, J., Stachura, Z., Hrynkiewicz, A.Z., 1999. structure and mechanics in fibroblasts: an atomic force microscopy study.
Elasticity of normal and cancerous human bladder cells studied by scanning Biophys. J. 78 (1), 520–535.
force microscopy. Eur. Biophys. J. 28 (4), 312–316. Sasaki, S., Morimoto, M., Haga, H., Kawabata, K., Ito, E., Ushiki, T., Abe, K.,
Lekka, M., Laidler, Ignacak, J., Labedz, M., Lekki, J., Struszczyk, H., Stachura, Sambongi, T., 1998. Elastic properties of living fibroblasts as imaged using
Z., Hrynkiewicz, A.Z., 2001. The effect of chitosan on stiffness and force modulation mode in atomic force microscopy. Arch. Histol. Cytol. 61
glycolytic activity of human bladder cells. Biochim. Biophys. Acta 1540 (1), 57–63.
(2), 127–136. Sato, H., Nagayama, K., Kataoka, N., Sasaki, M., Hane, K., 2000. Local
Liang, X., Mao, G., Simon Ng, K.Y., 2004. Probing small unilamellar EggPC mechanical properties measured by atomic force microscopy for cultured
vesicles on mica surface by atomic force microscopy. Colloids Surf. B: bovine endothelial cells exposed to shear stress. J. Biomech. 33 (1), 127–135.
Biointerfaces 34, 41–51. Sato, H., Kataoka, N., Kajiya, F., Katano, M., Takigawa, T., Masuda, T., 2004.
Lieber, S.C., Aubry, N., Pain, J., Diaz, G., Kim, S.-J., Vatner, S.F., 2004. Kinetic study on the elastic change of vascular endothelial cells on collagen
Aging increases stiffness of cardiac myocytes measured by atomic force matrices by atomic force microscopy. Colloids Surf. B: Biointerfaces 34 (2),
microscopy nanoindentation. Am. J. Physiol. Heart Circ. Physiol. 287, 141–146.
H645–H651. Scheffer, L., Bitler, A., Ben-Jacob, E., Korenstein, R., 2001. Atomic force
Mahaffy, R.E., Park, S., Gerde, E., Käs, J., Shih, S.K., 2004. Quantitative pulling: probing the local elasticity of the cell membrane. Eur. Biophys. J.
analysis of the viscoelastic properties of thin regions of fibroblasts using 30, 83–90.
atomic force microscopy. Biophys. J. 86, 1777–1793. Schrot, S., Weidenfeller, C., Schäffer, T.E., Robenek, H., Galla, H.J., 2005.
Mahaffy, R.E., Shih, C.K., MacKintosh, F.C., Käs, J., 2000. Scanning probe- Influence of hydrocortisone on the mechanical properties of the cerebral
based frequency-dependent microrheology of polymer gels and biological endothelium in vitro. Biophys. J. 89, 3904–3910.
cells. Phys. Rev. Lett. 85, 880–883. Shroff, S.G., Saner, D.R., Lal, R., 1995. Dynamic micromechanical properties
Mathur, A.B., Truskey, G.A., Reichert, W.M., 2000. Atomic force and total of cultured rat atrial myocytes measured by atomic force microscopy. Am. J.
internal reflection fluorescence microscopy for the study of force transmis- Physiol. Cell Physiol. 269, C286–C292.
sion in endothelial cells. Biophys. J. 78, 1725–1735. Simon, A., Cohen-Bouhacina, T., Porte, V.C., Aime, J.P., Amedee, J., Bareille,
Mathur, A.B., Collinsworth, A.M., Reichert, W.M., Kraus, W.E., Truskey, G.A., R., Baquey, C., 2003. Characterization of dynamic cellular adhesion of
2001. Endothelial, cardiac muscle and skeletal muscle exhibit different osteoblasts using atomic force microscopy. Cytometry A 54 (1), 36–47.
viscous and elastic properties as determined by atomic force microscopy. J. Sugawara, M., Ishida, Y., Wada, H., 2002. Local mechanical properties of
Biomech. 34, 1545–1553. guinea pig outer hair cells measured by atomic force microscopy. Hear. Res.
Miyazaki, H., Hayashi, K., 1999. Atomic force microscopic measurement of the 174 (1–2), 222–229.
mechanical properties of intact endothelial cells in fresh arteries. Med. Biol. Sugawara, M., Ishida, Y., Wada, H., 2004. Mechanical properties of sensory and
Eng. Comput. 37 (4), 530–536. supporting cells in the organ of Corti of the guinea pig cochlea—study by
Mozafari, M.R., Reed, C.J., Rostron, C., Hasirci, V., 2005. A review of scanning atomic force microscopy. Hear. Res. 192 (1–2), 57–64.
probe microscopy investigations of liposome–DNA complexes. J. Liposome Takai, E., Costa, K.D., Shaheen, A., Hung, C.T., Guo, X.E., 2005. Osteoblast
Res. 15, 93–107. elastic modulus measured by atomic force microscopy is substrate depen-
Mozhanova, A.A., Nurgazizov, N.I., Bukharaev, A.A., 2003. Local elastic dent. Annals Biomed. Eng. 33 (7), 963–971.
properties of biological materials studied by SFM. SPM-2003. In: Proceed- Takeuchi, M., Miyamoto, H., Sako, Y., Komizu, H., Kuzumi, A., 1998.
ings Nizhni Novgorod, March 2–5, pp. 266–267. Structure of the erythrocyte membrane skeleton as observed by atomic
Murakoshi, M., Yoshida, N., Iida, K., Kumano, S., Kobayashi, T., Wada, H., force microscopy. Biophys. J. 74, 2171–2183.
2006. Local mechanical properties of mouse outer hair cells: atomic force Timoshenko, S.P., Woinowsky-Krieger, S., 1970. Theory of Plates and Shells.
microscopic study. Auris Nasus Larynx 33 (2), 149–157. McGraw-Hill, New York.
Nagayama, M., Haga, H., Kawabata, K., 2001. Drastic change of local stiffness Tolomeo, J.F., Steele, C.R., Holley, M.C., 1996. Mechanical properties of the
distribution correlating to cell migration in living fibroblasts. Cell Motil. lateral cortex of mammalian auditory outer hair cells. Biophys. J. 71 (1),
Cytoskel. 50 (4), 173–179. 421–429.
Nowakowski, R., Luckham, P., Winlove, P., 2001. Imaging erythrocytes under Vinckier, A., Semenza, G., 1998. Measuring elasticity of biological materials by
physiological conditions by atomic force microscopy. Biochim. Biophys. atomic force microscopy. FEBS Lett. 430, 12–16.
Acta 1514, 170–176. Wada, H., Usukura, H., Katori, Y., Kakehata, S., Ikeda, K., Kobayashi, T., 2003.
Ohashi, T., Ishii, Y., Ishikawa, Y., Matsumoto, T., Sato, M., 2002. Experimental Relationship between the local stiffness of the outer hair cell along the cell
and numerical analyses of local mechanical properties measured by atomic axis and its ultrastructure observed by atomic force microscopy. Hear. Res.
force microscopy for sheared endothelial cells. BioMed. Mater. Eng. 12, 177 (1–2), 61–70.
319–327. Wu, H.W., Kuhn, T., Moy, V.N., 1998. Mechanical properties of L929 fibro-
Pesen, D., Hoh, J.H., 2005. Micromechanical architecture of the endothelial cell blasts measured by atomic force microscopy: effects of anticytoskeletal
cortex. Biophys. J. 88, 670–679. drugs and membrane crosslinking. Scanning 20 (5), 389–397.
Radmacher, M., Fritz, M., Kacher, C.M., Cleveland, J.P., Hansma, P.K., 1996. Yamane, Y., Shiga, H., Haga, H., Kawabata, K., Abe, K., Ito, E., 2000.
Measuring the viscoelastic properties of human platelets with the atomic Quantitative analyses of topography and elasticity of living and fixed
force microscope. Biophys. J. 70 (1), 556–567. astrocytes. J. Electron Microsc. (Tokyo) 49 (3), 463–471.
Rosenbluth, M.J., Lam, W.A., Fletcher, D.A., 2006. Force microscopy of Yoshikawa, Y., Yasuike, T., Yagi, A., Yamada, T., 1999. Transverse elasticity of
nonadherent cells: a comparison of leukemia cell deformability. Biophys. myofibrils of rabbit skeletal muscle studied by atomic force microscopy.
J. 90, 2994–3003. Biochem. Biophys. Res. Comm. 256, 13–19.
Rotsch, C., Braet, F., Wisse, E., Radmacher, M., 1997. AFM imaging and Zhang, S., Kraus, W.E., Truskey, G.A., 2004. Stretch-induced nitric oxide
elasticity measurements on living rat liver macrophages. Cell Biol. Int. 21 modulates mechanical properties of skeletal muscle cells. Am. J. Physiol.
(11), 685–696. Cell Physiol. 287, C292–C299.