Biomedical Engineering Reference
In-Depth Information
to be directly involved in the biorecognition process. On such a basis, the authors
have suggested a possible mechanism involving the formation of a dynamic complex
coupled to a rolling process in which both the specific and the nonspecific sequences
of DNA play some role. Jiang et al. have investigated the interaction between dif-
ferent DNA aptamers and the human immunoglobulin E, which plays a key role in
allergic response (Jiang et al., 2003). They developed an immobilization strategy
of the biomolecules, resulting in a rather high number of unbinding events (from
4 to 10) in each pull-off event. Under these conditions, the Poisson statistics pro-
vided a good description of the unbinding force distribution, allowing to single out
the contribution from the rupture of a single pair of interacting biomolecules (see
Chapter 4). They found an unbinding force of about 150 pN for a single specificinter-
action. The reliability and reproducibility of the results were demonstrated by repeat-
ing the experiments using different substrates and tips. Interestingly, they observed
that the unbinding force value can be reduced by slightly increasing the concentration
of NaCl; this is witnessing also the high sensitivity of DFS measurements to reveal
even subtle changes in the physiological conditions. They have also compared the
response of aptamers in competition with antibodies by carrying out control exper-
iments in which aptamers were substituted with immunoglobulin E antibody. The
finding that the affinity of aptamers to protein target matches or even surpasses that
of antibodies lend significant grounds to the higher potentialities of aptamers for
therapeutic and diagnostic applications.
Along the same direction, Basnar et al. have developed an approach exploiting
aptamers for detection of thrombin, a protein bearing a high relevance in heart dis-
eases (Bernhard et al., 2006). The interaction between a tip functionalized with
aptamers and a gold substrate covered with thrombin molecules has resulted in a
rather large number of rupture events (up to 15) during each retraction step. Applica-
tion of the Poisson statistics has provided an unbinding force for each pair of about
5 pN; such a value, which is close to the instrumental limits of the AFM equip-
ment, has been put into relationship to a melting of the quadruplex structure for the
aptamers. These results may also be rewarding in light of extending the effective
capability of DFS to the detection of extremely low interaction forces.
6.3.5 DFS
OF
C
OMPLEXES
I
NVOLVED IN
A
DHESION AND
A
GGREGATION
P
ROCESSES
A broad spectrum of biological processes requires controlled cell adhesion, that is,
the binding of a cell to a substrate, which can be another cell, a surface, or an organic
matrix. For example, cell-adhesion is involved in embryonic development, assem-
bly of tissues, cellular communication, inflammation and wound healing, tumour
metastasis, cell culturing, viral and bacterial infection, and so on. Cell adhesion is
commonly regulated by specific cell-adhesion molecules (CAMs), which are typ-
ically transmembrane receptors, and comprise an intracellular domain interacting
with cytoplasmic proteins and an extracellular domain that specifically binds to adhe-
sion partners (Kemler, 1992). The major classes of CAMs in mammals are cadherins,
selectins, integrins, and those belonging to the immunoglobulin superfamily. Some
results from DFS studies on systems involved in adhesion processes are reported
in Table 6.5.
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