Tomographic flow cytometry by digital holography

TLDR

High‑throughput single‑cell analysis is challenging, and while label‑free tomographic phase microscopy offers a promising route, in‑line tomography is difficult to implement because it requires complex sample rotation and multi‑direction probing. The study demonstrates that random rolling of cells in a microfluidic channel enables in‑line phase‑contrast tomography using smart wavefront analysis strategies. The method applies digital holography wavefront analysis to cells flowing in a microfluidic channel, as shown for red blood cells and diatom algae. The approach removes the need for a priori 3D position or orientation knowledge, allows continuous‑flow cytotomography without mechanical scanning, and accurately characterizes both red blood cells and diatom algae, indicating extensibility to diverse cell types.

Abstract

High-throughput single-cell analysis is a challenging task. Label-free tomographic phase microscopy is an excellent candidate to perform this task. However, in-line tomography is very difficult to implement in practice because it requires a complex set-up for rotating the sample and examining the cell along several directions. We demonstrate that by exploiting the random rolling of cells while they are flowing along a microfluidic channel, it is possible to obtain in-line phase-contrast tomography, if smart strategies for wavefront analysis are adopted. In fact, surprisingly, a priori knowledge of the three-dimensional position and orientation of rotating cells is no longer needed because this information can be completely retrieved through digital holography wavefront numerical analysis. This approach makes continuous-flow cytotomography suitable for practical operation in real-world, single-cell analysis and with a substantial simplification of the optical system; that is, no mechanical scanning or multi-direction probing is required. A demonstration is given for two completely different classes of biosamples: red blood cells and diatom algae. An accurate characterization of both types of cells is reported, despite their very different nature and material content, thus showing that the proposed method can be extended by adopting two alternate strategies of wavefront analysis to many classes of cells.