High-throughput multiparametric imaging flow cytometry: toward diffraction-limited sub-cellular detection and monitoring of sub-cellular processes
Gregor Holzner, Bogdan Mateescu, Daniël van Leeuwen, Gea Cereghetti, Reinhard Dechant, Stavros Stavrakis, Andrew J. deMello
Abstract
We present a sheathless, microfluidic imaging flow cytometer that incorporates stroboscopic illumination for blur-free fluorescence detection at ultra-high analytical throughput. The imaging platform is capable of multiparametric fluorescence quantification and sub-cellular localization of these structures down to 500 nm with microscopy image quality. We demonstrate the efficacy of the approach through the analysis and localization of P-bodies and stress granules in yeast and human cells using fluorescence and bright-field detection at analytical throughputs in excess of 60,000 and 400,000 cells/s, respectively. Results highlight the utility of our imaging flow cytometer in directly investigating phase-separated compartments within cellular environments and screening rare events at the sub-cellular level for a range of diagnostic applications.