Dynamic Real-Time Deformability Cytometry - Time-Resolved Mechanical Single Cell Analysis at 100 Cells/s

Abstract

In life sciences, the material properties of suspended cells have attained significance close to that of fluorescent markers but with the advantage of label-free and unbiased sample characterization. Until recently, cell rheological measurements were either limited by acquisition throughput, excessive post processing, or low-throughput real-time analysis. Real-time deformability cytometry, a label-free technique for single cell mechanical analysis with high-throughput of up to 1,000 cells / second, expanded the application of mechanical cell assays to fast on-the-fly phenotyping of large sample sizes, but has been restricted to single material parameters as the Young's modulus. Dynamic RT-DC (dRT-DC) overcomes this limitation and is capable to capture full viscoelastic properties of up to 100 suspended cells / second. Cellular shape-changes along the entire length of the microfluidic channel are tracked in real-time and are subsequently analyzed by a Fourier decomposition. We demonstrate that this approach allows to disentangle cell response to the complex hydrodynamic environment at the inlet from the steady-state stress distribution inside the channel. A superposition of both effects is present in almost all microfluidic systems and potentially biases label-free cytometric measurements relying on steady-state flow conditions. Dynamic RT-DC was performed on a precursor myeloid cell line HL60 as well as on primary blood cells. We show for the cell line that our method is capable to monitor changes in the apparent Young's modulus and viscosity after actin depolymerization. We use these findings for a first rheological comparison between erythrocytes, granulocytes and peripheral blood mononucleated cells (PBMCs) in a single experimental assay. We demonstrate that granulocytes and PBMCs have a significantly increased Young's modulus as well as viscosity compared to red blood cells and that our method is capable of a label-free discrimination of B- and CD4+ T-lymphocytes.