Biomedical Engineering Reference
In-Depth Information
where
C
b
and
l
rms,b
are the concentration and the rms of the end-to-end distance of
the back-filling molecules, respectively;
C
l
and
l
rms,l
are the concentration and the
rms of the end-to-end distance of polymeric linker that holds recognition partner
molecule, respectively; and
R
is the radius of the tip curvature. For example, if poly-
mer linker and the back-filling polymer have 40 and 15 statistical units, respectively,
with the statistical (Kuhn) length of 0.7 nm, and the probe with 40 nm radius of
curvature the dilution ratio of the linker to the back-filling polymer factor is
1:25.
For back-filling molecules significantly shorter than the linkers Equation 4.4 might
underestimate the dilution factor because of the entropy of mixing effects: Fraction
of longer polymeric molecules that chemically attach to surfaces is higher than the
fraction of longer polymers in solution (Al-Maawali et al. 2001). Therefore, Equation
4.4 provides only the starting point for experiments that aim to significantly reduce
multiple bond artifacts. It should be noted that the aspect of avoiding multiple inter-
actions remains poorly researched in DFS literature.
∼
4.6 CONCLUSION
In this chapter, we have considered methods of attaching molecules to surfaces in
DFS experiments primarily focusing on the most widely used approaches. The main
conclusion of comparing physical and chemical methods of attaching molecules
is that in spite of the simplicity of sample preparation using physical attachment
method, this method requires substantial verification for various artifacts. Chemical
methods of attaching molecules, particularly approaches that utilize polymeric teth-
ers, remove majority of limitations pertinent to the physical methods of attachment.
Chemical methods discussed in this chapter focus on approaches that have been
employed in DFS literature. Also, we have indicated that specific requirements of
DFS technique are considerably different from other techniques that employ immo-
bilized biomolecules. Commercially available cross-linking compounds and func-
tionalized tethers enable wide spread use of chemical attachment in DFS experi-
ments and facilitate application of this methodology by biophysics scientists who
do not have extensive chemistry training. Arsenal of available reactions to attach
molecules to interfaces is considerably more extensive than reactions discussed here.
It might be expected that newly developed reactions and experimental methodologies
will help solve the remaining problem of multiple interactions in DFS and facilitate
widespread use of DFS methodology as an analytical tool in biophysical research.
REFERENCES
Ahmad, S. F., L. A. Chtcheglova, B. Mayer, S. A. Kuznetsov, and P. Hinterdorfer. 2011.
Nanosensing of Fc gamma receptors on macrophages.
Analytical and Bioanalytical Chem-
istry
399 (7):2359-2367.
Al-Maawali, S., J. E. Bemis, B. B. Akhremitchev, R. Leecharoen, B. G. Janesko, and G. C.
Walker. 2001. Study of the polydispersity of grafted poly(dimethylsiloxane) surfaces using
single-molecule atomic force microscopy.
Journal of Physical Chemistry B
105 (18):3965-
3971.
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