HIGH-THROUGHPUT MICROARRAY-BASED MICRODIALYSIS (FAST-MD) WITH PARALLEL MEASUREMEN

DESCRIPTION (provided by applicant): Microdialysis is a versatile method for investigation of chemical compounds (analytes) in the extracellular microenvironment of a brain and other organs of the body. In spite of its tremendous importance in basic, clini cal and translational research, current approach has two major limitations. One of them is a low temporal resolution of measured analytes that is limited by the rate of diffusion and required sample volumes. Second drawback is the lack of direct coupling w ith the electrical processes that correspond to the concentration of a certain chemical compound. Therefore, there is a great need for an instrument or approach that would allow measuring the analytes close to real time permitting their analysis and correl ation in parallel with the changes in electrical activity. NeuroInDx proposes a novel method of nearly real time brain analyte measurements utilizing fluorescence generating enzymatic reactions and microarray-based sample collection and detection. In contr ast to the existing microdialysis approach, samples will be continuously deposited using a robotic arm on the surface of a polylysine coated glass slide that creates a microarray. After preprocessing, samples will be analyzed with a microarray scanner. Acq uired signal intensity data that is proportional to the specific analyte's concentration will be integrated with the original electrophysiological file using modified DataPac 2K2 software (RUN Technologies, Inc). Therefore, the correlation between brain el ectrical activity and the analyte's concentration can be established. In contrast to the existing biosensors, which can measure only one compound, Fast MD approach will allow to measure several compounds simultaneously. The long-term goal of NeuroInDx is t o develop and commercialize this Fast-MD device for clinical and basic research use. The main goal of the Phase I project is to demonstrate the feasibility of the Fast-MD approach for brain neurotransmitter analysis on the basis of the measurement of gluta mate, which is a major excitatory neurotransmitter in the brain, and glucose, a major correlate of brain metabolism. The proposed Fast-MD system can be used for the analysis of concentrations of different chemical compounds in the extracellular space of th e brain, muscle, lung, liver and other parts of the body. The experiments can be performed in the human during intensive care monitoring, as well as in freely moving and anesthetized animals with parallel wide frequency band recording of electrical activit y at the level of EEG, local field potentials and single neurons. PUBLIC HEALTH RELEVANCE: The approach utilizes continuous collection of cerebro-spinal fluid using microfluidics, analyte detection using enzymatic reaction that generates a fluorescent sign al, robotic deposition of the developed samples in a microarray format, and parallel monitoring of brain electrical activity. Phase I project is focused on the development of the Fast-MD prototype, its key components, process optimization and enzymatic det ection of glutamate and glucose.