Biological Applications of Dynamic Force Spectroscopy - Dynamic Force Spectroscopy and Biomolecular Recognition - page 227

et al., 2007). Such an approach has allowed Chtcheglova et al. to visualize, iden-
tify, and quantify local receptor binding sites at single-molecule level and to assign
their locations to the topographical features of the cell surfaces. Such a combined
approach is expected to become more and more followed in the study of biorecog-
nition processes since it offers the possibility to extract simultaneous dynamical and
spatial information on the systems at single-molecule level. Fritz et al. have inves-
tigated the interaction between P-selectin and P-selectin ligand (Fritz et al., 1998).
P-selectin belongs to a family of glycoproteins that mediate adhesion with leuko-
cytes or platelets and it is located within the endothelial cell wall, whereas the P-
selectin ligand is expressed in the microvilli at the leukocyte surface. In particular,
P-selectin supports leucocyte rolling under hydrodynamic flow via the interaction
with its ligand. Interestingly, the authors have hypothesized that the application of
the external force in the DFS experiments could mimic the physiological process. In
other words, the binding and unbinding processes occurring during the DFS experi-
ments could describe a rolling process on a surface under the action of a physiolog-
ical fluid as depicted in Figure 6.11. Under this assumption, they have determined
the dissociation rate from an analysis of the unbinding force trend versus the corre-
sponding unbinding length in terms of a modified freely jointed chain model with
a Monte Carlo simulation of the unbinding force distribution. They found a disso-
ciation rate of 0.022 s − 1 , which is indicative of a rather fast process, which could
be due to the high chain-like elasticity of the system. The dissociation rate was also
determined by SPR by finding a value of 3 10 − 4 s − 1 , which is markedly lower than

that obtained by DFS. In general, it has been assumed that the SPR values, obtained
without the application of external forces, could represent a lower limit for the DFS

·

(a)

Flow

Rolling
process

(b) Piezo displacement

DFS
experiment

FIGURE 6.11 (a) Sketch of ligand-receptor interaction during leukocyte rolling. (b) Sketch
of ligand-receptor unbinding during a DFS experiment. The retraction of the AFM cantilever
from the substrate well mimics the physiological rolling process at the origin of the unbinding.
(Adapted from Fritz, J. et al. 1998. Proc. Natl. Acad Sci. U.S.A. , 95:12283-12288.)