Biomedical Engineering Reference
In-Depth Information
the steady-state position of labeled components in a lowing stream, the concentration of very
dilute (<1 nM) analytes can be measured in a few microliters of sample in seconds. his assay
has been demonstrated in the format of a small molecule analyte competition immunoassay
using luorescence imaging detection, and for larger analytes as well. he DIA can also monitor
concentrations of analytes as large as proteins, at the cost of increased assay times. Most excit-
ing is that this assay can operate almost completely without sample preconditioning, greatly
strengthening its applicability in point-of-care diagnostics.
4.5.3 Heterogeneous Phase (Surface-Bound) Immunoassays
he utilization of the walls of a microchannel allows access to the greater complexity and sensi-
tivity of assays like ELISA. his strategy requires very good control of surface binding (
k
on
and
k
of
), as illustrated in
Figure 4.11
, because mass transport to and from the surface is afected by
difusion (a “force” that works isotropically), convection (which works in the direction of low),
and the presence of electrical ields (if any).
he irst microluidic immunoassay, consisting of a set of PDMS channels that delivered anti-
bodies to micron-sized lines of a glass substrate for luorescence detection, dates from 1997
and is credited to Hans Biebuyck's group (then at IBM Zurich), as explained in Section 2.4.3
(see
Figure 2.17
). Recent implementations of immunoassays using surface plasmon resonance
detection have also been made in microluidic implementations.
Microluidics is generally appealing because it provides low-cost automation solutions, so it is
slowly becoming the technological substitute of luidic robotics. For a much lower price, it also
consumes fewer reagents. (he main drawbacks are that it tends to operate on nonstandard plat-
forms and requires some uncommon expertise, but those barriers are being lowered every day.)
A very common problem in immunoassays is to establish a serial dilution curve to test anti-
body binding response. hese dilutions are cumbersome to produce, requiring several pipetting
steps with traditional multiwell pipettors. Several microluidic titrators have been developed
(see Section 3.9.2). As an example,
Figure 4.12
shows a serial dilution luorescent immuno-
assay device reported in 2003 by George Whitesides' group from Harvard University. he assay
can measure the concentrations of multiple antibodies in parallel in one experiment using a
network of microchannels to achieve serial dilution. he branching structures of the microlu-
idic network serially dilute one stream by half by connecting it to the adjacent stream (so long
as proper mixing occurs at each stage, which is ensured with the incorporation of “chevron”
grooved mixers on the surface of the microchannels). To illustrate the assay, the concentration
of antibodies in HIV + human serum (anti-gp41 and anti-gp120) was determined by secondary
Diffusion
Convection
Supply
Electrophoresis
k
on
k
off
Site occupancy
FIGURE 4.11
Mass.transport.in.microluidic.immunoassays..(Figure.contributed.by.Paul.Yager.)
