Super-multiplexed vibrational imaging
Modern science and technology have exploding demands for multiplexing techniques to simultaneously measure a large number of distinctive species. Notable scientific research areas include exploring structure–function relationships in brain, in situ RNA profiling for spatial transcriptome; understanding heterogeneity of tumor microenvironments; revealing intricate organelle interactions of living cells. Meanwhile, multiplexing biotechnology applications include cell phenotyping, sorting and tracing, high-throughput detection of antigens for diagnostics, and high-throughput screening for drug discovery. All these multiplexing-central areas are beyond the reach of fluorescence microscopy. Therefore, transformative imaging technology is urgently needed to overcome the spectral limitation and break this longstanding “color barrier”.
Recently we conceived a novel optical super-multiplex imaging platform. Our invention consist of two components: (1) introducing a new paradigm of coherent Raman microscopy; (2) creating a matching palette of imaging probes with sharp (~15 cm-1) Raman peaks. Physically, we have developed electronic pre-resonance stimulated Raman scattering (epr-SRS) microscopy, achieving sensitivity down to several tens of molecules while retaining exquisite vibrational selectivity. Hence, epr-SRS microscopy can be complementary to or even supersede fluorescence in multiplex applications. Chemically, we have designed and synthesized a new palette of Raman probes (named MARS dyes) that enabled simultaneous 24-color imaging with potential for expansion. MARS dyes exhibit 100 times sharper Raman peaks than fluorescence spectra. Biologically, we demonstrated its utility in various cells and tissues, monitored proteome turnover dynamics, and discovered metabolic heterogeneity between neuron and astrocytes.
L. Wei, Z. Chen, L. Shi, R. Long, A. V. Anzalone, L. Zhang, F. Hu, R. Yuste, V. W. Cornish and W. Min. “Super-multiplex vibrational imaging”, Nature, 544, 465 (2017).
F. Hu, C. Zeng, R. Long, Y. Miao, L. Wei, Q. Xu and W. Min. Super-multiplexed optical imaging and barcoding with engineered polyynes. Nature Methods (2018) in press