Single-Cell Biophysics: Measurement, Modulation, and Modeling

Single-Cell Biophysics: Measurement, Modulation, and Modeling

Poster Abstracts

74-POS Board 37 Deformability Cytometry and 1D Fluorescence Imaging in Real-Time

Philipp Rosendahl 1 , Katarzyna Plak 1 , Angela Jacobi 1 , Nicole Töpfner 1,2 , Jochen Guck 1 . 1 Biotechnology Center, Technische Universität Dresden, Dresden, Germany, 2 University Clinic Carl Gustav Carus, Technische Universität Dresden, Dresden, Germany. During the last decades, tools for rapid characterization of large cell quantities have become indispensable not only for basic research but also clinical diagnostics. The gold standard for cell characterization is flow cytometry. Its success is closely tied to the availability of fluorescent labels. But what if there is no molecular marker known for the cells of interest? Or if the label changes cell function? Or cells shall be used for transplantation? As an attractive alternative, deformability cytometry exploits cell mechanics as a sensitive, inherent, label-free functional marker, but lacks the specificity provided by a fluorescent signal. Here we present real-time fluorescence deformability cytometry (RT-FDC), the ideal, combined system. It facilitates fluorescence detection as in conventional flow cytometry, extended by 1D analysis of spatial information encoded in the fluorescence pulse shape, and adds bright field imaging for mechanical phenotyping of single cells — all in real-time at rates of 100 cells/s. We show utility of RT-FDC for the most common fluorescent labels: Fluorescent surface markers (CD34) are used to separate human hematopoietic stem and progenitor cells (HSPCs) from an unpurified apheresis sample as harvested for bone marrow transplantations. Membrane permeant dyes identify reticulocytes by their ribonucleic acid (RNA) content in a blood sample. And endogenously expressed fluorescent proteins (FUCCI) reveal cell cycle phases in an unsynchronized sample of retinal pigment epithelial cells (RPE1). In addition, we can now also directly correlate mechanical characteristics with fluorescence intensity and localization for each single cell to improve correct classification. In future, this combined approach could establish mechanical phenotyping as equivalent to fluorescent labeling, or even identify subpopulations invisible to molecular labels. RT-FDC is destined to find wide-spread utilization and will help to further improve the applicability of flow cytometry.

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