A unique magnetic bead retention mechanism has been developed by Lacharme
et al., [ 46 ] which exploited the self-assembling properties of magnetic beads that
led to the formation of a magnetic chain, without the use of sophisticated fluidic
handling. The magnetic chain was formed by trapping magnetic beads in
microchannels with cross-sections with periodically enlarged size. When a mag-
netic field is applied, the magnetic beads self-assembled as chains along the length
of the channel. This mechanism provided efficient mixing and as a result enhanced
the antigen capture in an immunoassay performed completely on-chip with detec-
tion limit in the range of a few ng/mL in 30 min, using only nL of reagents and
antibody solution.
2.3.2 Non-magnetic Microbeads
Non-magnetic microbeads require physical retention for manipulation, and the
removal of unbound analyte. Physical retention of non-magnetic microbeads can
be achieved by microstructures fabricated within the microfluidic system. The
Kitamori group used a dam structure to trap antibody-coated polystyrene beads
[ 47 , 48 ]. Ko et al. [ 49 ] developed an integrated chip for protein sensing using
antibody-coated polystyrene beads trapped in a reaction area by micropillars made
of PDMS. Sample solution, gold nanoparticles labeling agent, and signal generation
solution were then flow through the trapped beads. The gold nanoparticles cata-
lyzed the silver ions in the signal generation solution into metallic silver, which in
turn deposited onto the gold nanoparticles and thereby enlarged the particles to
100 nm. The enlarged particles formed an electrical bridge between the electrodes
layered on the sensing surface, resulting in a signal due to the decrease in resistance.
The entire chip-based assay can be completed in less than an hour, while
performing the same assay with a conventional setup requires at least several
hours on hands on time.
Employing a configuration similar to solid-phase extraction, Shin et al. [ 50 ]
packed antibody-conjugated microbeads against a frit structure as a strategy to
increase the sensitivity of immunoassay for C-reactive protein (CRP). Captured
CRP were captured in the packed bead matrix, and subsequently eluted in an acidic
buffer. The extraction also worked successfully to detect CRP in complex sample
such as serum and cerebrospinal fluid [ 51 ].
Non-magnetic microbeads can also be immobilized by dielectrophoresis [ 52 ]or
electrostatic forces [ 53 , 54 ] without the use of microstructures for physical retention
of the beads. Holmes et al. [ 52 ] developed a microchip-based flow cytometer using
commercial methacrylate beads for capture antibody immobilization. The
microchip-based flow cytometer used negative dielectrophoresis to focus the
microbeads into a stream of particle, and the impedance and fluorescence signals
of each bead were measured.
In the work by Sivagananam, [ 53 ] positively charged APTES were patterned on
a glass substrate, and negatively charged streptavidin-coated microbeads were
assembled electrostatically. The assembled beads were then used for conjugation
9 On-Chip Immunoassay for Molecular Analysis 229