Smartphone Imaging Flow Cytometry for High-Throughput Single-Cell Analysis.

We present a portable imaging flow cytometer comprising a smartphone, a small-footprint optical framework, and a PDMS-based microfluidic device. Flow cytometric analysis is performed in a sheathless manner via elasto-inertial focusing with a custom-written Android program, integrating a graphical user interface (GUI) that provides a high degree of user control over image acquisition. The proposed system offers two different operational modes. First, "post-processing" mode enables particle/cell sizing at throughputs of up to 67 000 particles/s. Alternatively, "real-time" mode allows for integrated cell/particle classification with machine learning at throughputs of 100 particles/s. To showcase the efficacy of our platform, polystyrene particles are accurately enumerated within heterogeneous populations using the post-processing mode. In real-time mode, an open-source machine learning algorithm is deployed within a custom-developed Android application to classify samples containing cells of similar size but with different morphologies. The flow cytometer can extract high-resolution bright-field images with a spatial resolution <700 nm using the developed machine learning-based algorithm, achieving classification accuracies of 97% and 93% for Jurkat and EL4 cells, respectively. Our results confirm that the smartphone imaging flow cytometer (sIFC) is capable of both enumerating single particles in flow and identifying morphological features with high resolution and minimal hardware.

Ultrahigh-Throughput, Real-Time Flow Cytometry for Rare Cell Quantification from Whole Blood.

We present an ultrahigh-throughput, real-time fluorescence cytometer comprising a viscoelastic microfluidic system and a complementary metal-oxide-semiconductor (CMOS) linear image sensor-based detection system. The flow cytometer allows for real-time quantification of a variety of fluorescence species, including micrometer-sized particles and cells, at analytical throughputs in excess of 400,000 species per second. The platform integrates a custom C++ control program and graphical user interface (GUI) to allow for the processing of raw signals, adjustment of processing parameters, and display of fluorescence intensity histograms in real time. To demonstrate the efficacy of the platform for rare event detection and its utility as a basic clinical tool, we measure and quantify patient-derived circulating tumor cells (CTCs) in peripheral blood, realizing that detection has a sensitivity of 6 CTCs per million blood cells (0.000006%) with a volumetric throughput of over 3 mL/min.

Oscillatory Viscoelastic Microfluidics for Efficient Focusing and Separation of Nanoscale Species.

The ability to precisely control particle migration within microfluidic systems is essential for focusing, separating, counting, and detecting a wide range of biological species. To date, viscoelastic microfluidic systems have primarily been applied to the focusing, separation, and isolation of micrometer-sized species, with their use in nanoparticle manipulations being underdeveloped and underexplored, due to issues related to nanoparticle diffusivity and a need for extended channel lengths. To overcome such issues, we herein present sheathless oscillatory viscoelastic microfluidics as a method for focusing and separating both micrometer and sub-micrometer species. To highlight the efficacy of our approach, we segment our study into three size regimes, namely, micrometer (where characteristic particle dimensions are above 1 µm), sub-micrometer (where characteristic dimensions are between 1 µm and 100 nm), and nano (where characteristic dimensions are below 100 nm) regimes. Based on the ability to successfully manipulate particles in all these regimes, we demonstrate the successful isolation of p-bodies from biofluids (in the micrometer regime), the focusing of λ-DNA (in the sub-micrometer regime), and the focusing of extracellular vesicles (in the nanoregime). Finally, we characterize the physics underlying viscoelastic microflows using a dimensionless number that relates the lateral velocity (due to elastic effects) to the diffusion constant of the species within the viscoelastic carrier fluid. Based on the ability to precisely manipulate species in all three regimes, we expect that sheathless oscillatory viscoelastic microfluidics may be used to good effect in a range of biological and life science applications.