Progress has been made toward meeting several technical challenges in a continuing effort to develop “smart dust” sensors for remote detection of chemical and biological agents. Until now, the state of the art of laboratory-on-a-chip devices has been typified by microfluidic cartridges variously mounted in or connected to desktop or handheld devices. Smart dust sensors would be smaller: they would be submillimeter-sized devices, the implementation of which would necessitate levels of miniaturization and integration heretofore known only in biological systems.
A promising approach toward the development of highly functional smart dust prototypes involves the design and fabrication of the prototypes as hybrid devices, in which biological nanomachines would be integrated into microfabricated synthetic structures. A representative example of a smart dust biosensor according to this approach is depicted schematically in the figure. In this example, antibody functionalized microtubules would capture analytes [that is, target chemical or biological agents (e.g., target viruses)] in a first chamber, molecular shuttles driven by biomolecular motors of which the microtubules would be integral parts would transport the analytes to a second chamber wherein the analytes would be tagged with quantum dots or fluorescent-dye-containing microspheres, then the shuttle motors would transport the tagged analytes to a third chamber, wherein the tags (and, hence, the analytes) would be detected remotely by fluorescence measurements.
Major technical challenges involved in this approach and the progress made in this development effort toward meeting the challenges are summarized as follows:
- Temperature stabilization of biomolecular motor powered devices - The activity rates of natural biological nanomachines vary significantly with temperature, making it necessary to regulate temperatures in traditional biotechnological and microfluidic devices. However, regulation of temperature would not be feasible in smart dust sensors, making it desirable to make activity rates remain as nearly constant as possible in the face of temperature changes. In this effort, experiments were performed on motor proteins, which are central in a number of nanobiodevices currently under development. The results of the experiments led to the conclusion that provided that certain preconditions are satisfied, at least partial temperature stabilization of enzyme activity levels can be achieved at the expense of enzyme turnover by reducing substrate concentrations substantially.
- Measurement of brightnesses of fluorescent tags for remote detection. - The brightnesses of fluorescent tags are critical design parameters. In this effort, absolute brightnesses of samples of fluorescent- dye-containing micro- spheres and variations of relative brightness among individual microspheres were measured.
- Optimization of velocity of the molecular shuttles for pick-up of fluorescent tags in the second chamber. - The need for optimization of velocity of molecular shuttles was demonstrated in experiments on the attachment of nanospheres via biotin-streptavidin linkages. The optimum speed depends on the characteristic times of subprocesses involved in the formation of the linkages. In essence, biotin and streptavidin act as glue that must harden for a sufficient time before stress can be applied.
- Proof-of-principle demonstration of a basic biomolecular motor powered sensor. - A circular well structure was fabricated by use of a photoresist. Biotinylated microtubules and biotinylated fluorescent microspheres were deposited in a central region of the circular well, leaving a region at the perimeter that was initially free of fluorescent micro-spheres. Injection of streptavidin (as a model of analyte molecules) and subsequent photolytic release of caged adenosine triphosphate led to capture of streptavidin by microtubules, pick-up of microspheres, and, finally, immobilization of microsphere-carrying microtubules at the wall of the well. This shows that molecular shuttles can drive the analyte-dependent transport of fluorescent tags.
This work was done by Henry Hess of the University of Florida for the Air Force Research Laboratory.
This Brief includes a Technical Support Package (TSP).
Advances Toward Development of "Smart Dust" Biosensors
(reference AFRL-0070) is currently available for download from the TSP library.
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