TRD 1: FLUORESCENCE SPECTROSCOPY AND MICROSCOPY TECHNIQUES Investigators: P. So (1.1, 1.2) [lead]; M. Bawendi (1.2); G. Schlau-cohen (1.3) Collaborative Projects: Jain (CP1), Boyden (CP4), Campagnola (CP8), Coleman (CP9) Project Summary: Fluorescence spectroscopy and imaging are key techniques in the repertoire of the biomedical research community. In the LBRC, the investigators leverage their expertise in precision spectroscopy, contrast agent development, and coherent spatial and temporal control of ultrafast pulses to develop cutting-edge technologies for analyte-specific investigation of biological systems, from proteins to whole organisms. This fluorescence-based TRD builds upon 3D light sculpting techniques and short-wave infrared (SWIR) technologies developed in the current cycle with three exciting new directions: high-throughput deep SWIR imaging (TRD1.1), high-throughput, super-resolution 3D imaging (TRD1.2), and the nanometer- scale study of protein motions (TRD1.3). These directions are motivated by LBRC collaborations. Pushed by the study of cancer biology inside thick solid tumors in vivo, especially for monitoring dynamic events like blood flow and variations in oxygenation (CP1), TRD1.1 seeks to optimize both imaging speed and depth by combining patterned two-photon temporally focused wide-field excitation with compressive-sensing algorithms to image ultra-bright quantum dots (TRD4). Pushed also by Dr. Boyden's work to map the connection diagram of the brain (CP4), which in turn requires high-throughput identification of synaptic clefts at 50 nm resolution throughout a 0.5 cm3 volume. Based on our expertise in structured illumination (SI) and point spread function (PSF) engineering, TRD1.2 seeks to improve super-resolution imaging speed to approach 1G voxel/sec in order to map the whole brain within ~1 year. The same super-resolution approach is employed for high- throughput 3D microfabrication of an extracellular matrix to control cancer cell migration and tissue regeneration (CP8). Finally, pushed by the need for new insight into the signaling mechanisms of receptors, which are the targets of cancer therapeutics (CP9), TRD1.3 will develop fluorescence spectroscopy tools with nanometer spatial and sub-millisecond temporal resolution. In summary, this TRD further extends the core strength of the LBRC in fluorescence instrumentation by introducing these three new research directions.
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