Biomedical Engineering Reference
In-Depth Information
Haes et al. (2006)
recently reported that detection techniques for target proteins are in
demand; especially those that are fast, sensitive, selective, and without interference from
background materials. Immunoassays are widely used bioanalytical methods (
Elkins, 1989;
Diamindis and Christpoulos, 1996
).
Haes et al. (2006)
point out that all immunoassays use
antibodies as capture molecules for strong and specific binding to target antigens. They fur-
ther explain that immunoassay techniques have been miniaturized into microchip platforms
(
Sato et al., 2002, 2004; Gao et al., 2005; Rubina et al., 2005; Herr et al., 2005; Phillips
and Cheng, 2005
).
Haes et al. (2006)
carefully delineate the parameters that are involved in immunoassay tec-
hniques. These include assay simplicity, convenience, cost, total assay time, assay sensitivity,
and reagent stability. These authors also point out the limitations of immunoassay techniques
which include limitations in the binding rate coefficient in the antigen-antibody interaction,
the presence of non-specific binding, the diffusion distance between the antigen (e.g., in solu-
tion) and the antibody (e.g., immobilized on the sensor surface), detector sensitivity, and
assay time. These authors have developed a displacement immunoassay for the detec-
tion of SEB on a glass microchip. They used laser-induced fluorescence detection and
electrokinetically controlled fluidic delivery. The authors further point out that a monoclonal
antibody that is specific for SEB is covalently attached to the microbeads. These beads were
trapped on a microchip using narrow pathways on custom-made microchip devices. The
authors emphasize that an apparent field enhancement enrichment phenomenon was obtained
due to varying field strengths within the nonuniform channels.
Haes et al. (2006)
analyzed the binding and dissociation of 100 aM (attamols) to 100 nM SEB
in solution to the antibody-functionalized microbeads on a microchip. The size of the beads
had two constraints: minimizing back pressure and the time required for the immunoassay.
The back pressure was minimized by minimizing the bead length in the microbeads. The second
constraint was satisfied by making sure that the antibody-coated beads attain saturation in
a “short” time scale. These authors noted that a bead length of around 200
m
m satisfied
both of the above constraints. Also, excellent sensitivity and dynamic range was attained.
Figure 14.8a
shows the binding and dissociation of 1 nM SEB in solution to the antibody-
functionalized microbeads on a sensor chip. The monoclonal antibody that is specific for
SEB is covalently attached to the silica beads. A dual-fractal analysis is required to describe
the binding and the dissociation kinetics. The values of (a) the binding rate coefficient,
k
, and
the fractal dimension,
D
f
, for a single-fractal analysis, (b) the binding rate coefficients,
k
1
and
k
2
, and the fractal dimensions,
D
f1
and
D
f2
, for a dual-fractal analysis, (c) the dissociation rate
coefficient,
k
d
, and the fractal dimension,
D
fd
, for a single-fractal analysis, and (d) the
dissociation rate coefficients,
k
d1
and
k
d2
, and the fractal dimensions,
D
fd1
and
D
fd2
, for a
dual-fractal analysis are given in
Tables 14.6
and
14.7
. The zero value obtained for the fractal
dimension,
D
f1
, for a dual-fractal analysis indicates that for this initial phase of binding the
