Biomedical Engineering Reference
In-Depth Information
Under certain circumstances after a protein has adsorbed to a particle
surface, lateral diffusion is known to occur.
29
This allows more of the NMs to
attach to a bigger protein molecule or vice versa.
4.3 IMMOBILIZATION STRATEGIES
Immobilization of probes in NM-enabled biosensors requires strategies that protect
the integrity of the NMs and the biomolecular probes. Proper immobilization tech-
niques help maintain the bioactivity via control of the orientation and conformation
of biomolecules. Retention of the active conformations of the biomolecules directly
affects both the biosensors stability and reproducibility of signals.
7,12,32-35
Bio-
molecules such as proteins,
36-38
DNA,
32,33,37,39,40
and whole cells,
11,12,33,37,39,41,42
directly adsorb on various surfaces rendering the activity of the targeted biomol-
ecules or portions of the biomolecules inactive.
39,43,44
Hence, a linker that protects
the biomolecule from direct exposure to sensor surfaces is usually employed for
attachment or as entrapment moiety
36,45-47
or a covalent attachment on functional
groups is usually employed as the entrapment agent.
42,48
With the emergence of
NMs, electrochemical biosensing with alternative ways for immobilization of
biomolecule recognition probes are now being developed.
42
NMs possess a high
surface-to-volume ratio and their network assemblies exhibit porous structure that
is a large area of active surface that alleviates surface-fouling effects that causes
the enhanced retention of bioactivity. This leads to a large amount of biological
molecules (e.g. enzyme, antibody, cell, and DNA) that can be immobilized on the
active surface of NMs that allows more intense response signal.
48
4.3.1 Direct Adsorption
Direct adsorption is one of the ways by which NMs are used in biosensors
development. Direct adsorption had been used in developing NM-based bio-
sensors for the detection of DNA,
49,50
proteins,
42,51-54
whole cells,
12,39,50,55,56
and parts of cells.
39,54,57
The group of Ehrenberg et al.
54
determined the time course of protein
adsorption through the incubation of -COOH functionalized NMs in serum.
Protein adsorption on the NMs surface took place in seconds and reached a
plateau in less than 5 min. Steady amounts of protein were adsorbed in less than
5 min indicating rapid adsorption compared with cellular binding. This relative
speed was also reported in the literature demonstrating adsorption rate constants
on the order of seconds
53
compared to timescales of at least minutes for attach-
ment to cell surfaces.
One of the materials to which many different kinds of molecules adsorb is
gold. AuNPs have found various applications in medicine. Biomolecules such
as single-stranded DNA (ssDNA) and double-stranded DNA (dsDNA) can be
nonspecifically adsorbed on AuNPs. Double- and single-stranded oligonucle-
otides (ONTs) have different electrostatic properties that arise as a result of
