Biomedical Engineering Reference
In-Depth Information
the protein of interest from the sample prior to sensor immobilization using protein
purification techniques, it is possible to increase capture protein density, thereby
maximizing signal.
7.4.2
Wash-Free Protein Detection Assay for POC
With magnetic biosensors, the traditional sandwich assay described above can
be redesigned to leverage the proximity-based detection capabilities and unique
magnetic properties of a magnetically responsive biosensor system. The GMR
nanosensors, for example, can be built with an ultrathin passivation layer. As
discussed above, these GMR sensors are proximity-based sensors, and therefore,
only magnetic nanotags within
150 nm of the surface are detected [
5
]. Because
the magnetic nanotags typically employed are on the order of 50 nm in diameter and
colloidally stable, they do not settle or precipitate on the sensor surface, contributing
negligible signal in the absence of the target protein or detection antibody. Only
in the presence of both the biomolecule of interest and detection antibody will the
magnetic nanotags congregate over the appropriate sensor in close enough proximity
and in high enough density for the GMR sensor to experience a measurable
magnetoresistance change. This is a significant advantage over the vast majority of
protein detection platforms in which the excess/unreacted tags must be washed away
prior to detection, preventing their ability to utilize a wash-free detection method.
As a result, while performing a traditional sandwich assay requires washing steps
to remove excess antibodies or nanotags, with magnetic nanotechnology, the signal
remains unchanged with a final wash step. Accordingly, the protein content can be
determined in the assay without implementing washing (Fig.
7.5
). Thus, because this
assay obviates the need for washing steps, it offers a faster, simpler testing process
that untrained users can easily perform in point-of-care settings.
By taking advantage of the “autoassembly” nature of this assay, only minimal
human intervention is required to run a test, removing the dependence on the end
user to have prior laboratory training. Moreover, the assay can be run in an open-well
format, removing the need for complex microfluidic plumbing or external pneumatic
pressure controllers. The wash-free assay entails only three steps: (1) The operator
places the biological sample into the reaction well which is equipped with an array
of GMR sensors pre-functionalized with a panel of antibodies against predetermined
proteins of interest. As the sample incubates in the well, the proteins of interest are
captured by the immobilized antibodies directly over individually addressable GMR
sensors. (2) The user adds a solution of magnetic nanotags labeled with streptavidin.
At this point, no detectable reaction takes place because no biotin is present in the
reaction well (Fig.
7.8
a). (3) Finally, detection antibodies labeled with biotin are
introduced. These detection antibodies subsequently link the magnetic nanotags to
the captured analyte, thus inducing a measurable signal in the underlying GMR
