Hasselbrink This research will investigate the use of biomolecular motors as the basis for a completely novel, molecule-specific transport mechanism that could have powerful applications in biomolecule sensors. This research combines biomolecular motor transport with an immosensing strategy that literally allows one to program sensor molecules to grab specific biomolecules from a liquid solution, and then carry them to a fixed detection window. The operation of such a sensor can be summarized as follows: a liquid sample is placed in a reservoir containing a solution of microtubules derivatized with antibodies, plus ATP, an oxygen scavenger. Labeled (or enzymelinked, or bead-bound) antibodies are then added. Fast (non- diffusion limited) and highly specific "sandwich"-type binding between microtubules, target analyte, and labeled antibody occurs, and a plunger is placed atop the reservoir and pressed to inject of this mixture into a cavity in a microfluidic chip. Within this cavity, the microtubules will translocate autonomously (using ATP omnipresent in the solution as fuel), but in a programmed direction towards a detection window on the chip, where they will concentrate for highly sensitive detection by LED/photodiode fluorescence. Microtubule directional programming is achieved using a patterned film of CytopTM (spin-on fluoropolymer) on the microfluidic chip inner surfaces. The film works by both allowing for selective patterning of kinesin, and by providing mechanical guides that direct the microtubules along the desired path of translocation. Proof-of-principle experiments have been performed, and demonstrated a rudimentary sensor based on this concept. Numerous questions regarding the specificity, speed, and sensitivity of sensors exploiting these phenomena remain to be answered.
The research plan is to test the generality of this microtubule-based immunoassay approach to a variety of antibodies, and to characterize the efficiency and speed of this approach in a variety of microdevices. Statistical studies will characterize and optimize translocation paths using image processing-based "particle tracking" studies of microtubule translocation. Concentrator designs will also be studied and optimized so that the highest possible signal may be obtained from simple, integrated LED/photodiode detection systems in the shortest possible time. Experiments on the robustness of microtubules under fluid shear, and dependencies of sensor performance on poorly-controlled variables such as temperature, will also be conducted.
