An automated platform for kinase assays on patient cells

点击翻译按钮获取中文摘要

DESCRIPTION (provided by applicant): An integrated laser microbeam-microscope system with time resolved imaging capabilities is proposed for studying laser induced tissue injury. Laser microbeams offer a fast, automated and non-contact means for cellular micromanipulation due to their ability to deposit energy with high spatial specificity and limited collateral damage. There is growing interest in their use for applications in tissue microprocessing and single cell analysis. However, research on basic mechanisms of laser-cell interactions is limited due to the small spatial scales and fast time scales over which they occur. The experimental platform we propose will image the dynamics of laser microbeam-induced injury with high spatial and temporal resolution in tissue engineered samples that mimic in-vivo systems. Focused nanosecond laser pulses will be delivered to the sample to cause injury at specific sites and the process will be imaged using time-resolved imaging on nanosecond to microsecond timescales. The contributions of the high-temperature plasma, Shockwave propagation and cavitation bubble expansion and collapse to cell injury will be determined from these images. Time-resolved images will also be used for quantitative estimation of cavitation bubble parameters such as bubble size, collapse time and bubble energy. The bubble dynamics will be used to estimate fluid velocity and shear stress at the tissue site. The biological effects of laser pulses will be monitored using fluorescence assays. A major goal would be to relate the physical effects from time-resolved imaging to the observed biological response. The hydrodynamic modeling will provide insights on cellular response to high shear fields on fast time scales to provide a biophysical characterization of the damage process. The platform will be designed to be scalable for general studies of laser microbeams and cavitation phenomena in biology. This research will be done primarily in Bangalore, India at the National Centre for Biological Sciences in collaboration with Dr. Kaustubh Rau as an extension of NIH grant R01-EB004436.