High-throughput viscoelastic characterization of cells in hyperbolic microchannels.

Extensive research has demonstrated the potential of cell viscoelastic properties as intrinsic indicators of cell state, functionality, and disease. For this, several microfluidic techniques have been developed to measure cell viscoelasticity with high-throughput. However, current microchannel designs introduce complex stress distributions on cells, leading to inaccuracies in determining the stress-strain relationship and, consequently, the viscoelastic properties. Here, we introduce a novel approach using hyperbolic microchannels that enable precise measurements under a constant extensional stress and offer a straightforward stress-strain relationship, while operating at a measurement rate of up to 100 cells per second. We quantified the stresses acting in the channels using mechanical calibration particles made from polyacrylamide (PAAm) and found that the measurement buffer, a solution of methyl cellulose and phosphate buffered saline, shows strain-thickening following a power law up to 200 s-1. By measuring oil droplets with varying viscosities, we successfully detected changes in the relaxation times of the droplets and our approach could be used to get the interfacial tension and viscosity of liquid-liquid droplet systems from the same measurement. We further applied this methodology to PAAm microgel beads, demonstrating the accurate recovery of Young's moduli and the near-ideal elastic behavior of the beads. To explore the influence of altered cell viscoelasticity, we treated HL60 human leukemia cells with latrunculin B and nocodazole, resulting in clear changes in cell stiffness while relaxation times were only minimally affected. In conclusion, our approach offers a streamlined and time-efficient solution for assessing the viscoelastic properties of large cell populations and other microscale soft particles.

Image-based cell sorting using focused travelling surface acoustic waves.

Sorting cells is an essential primary step in many biological and clinical applications such as high-throughput drug screening, cancer research and cell transplantation. Cell sorting based on their mechanical properties has long been considered as a promising label-free biomarker that could revolutionize the isolation of cells from heterogeneous populations. Recent advances in microfluidic image-based cell analysis combined with subsequent label-free sorting by on-chip actuators demonstrated the possibility of sorting cells based on their physical properties. However, the high purity of sorting is achieved at the expense of a sorting rate that lags behind the analysis throughput. Furthermore, stable and reliable system operation is an important feature in enabling the sorting of small cell fractions from a concentrated heterogeneous population. Here, we present a label-free cell sorting method, based on the use of focused travelling surface acoustic wave (FTSAW) in combination with real-time deformability cytometry (RT-DC). We demonstrate the flexibility and applicability of the method by sorting distinct blood cell types, cell lines and particles based on different physical parameters. Finally, we present a new strategy to sort cells based on their mechanical properties. Our system enables the sorting of up to 400 particles per s. Sorting is therefore possible at high cell concentrations (up to 36 million per ml) while retaining high purity (>92%) for cells with diverse sizes and mechanical properties moving in a highly viscous buffer. Sorting of small cell fraction from a heterogeneous population prepared by processing of small sample volume (10 µl) is also possible and here demonstrated by the 667-fold enrichment of white blood cells (WBCs) from raw diluted whole blood in a continuous 10-hour sorting experiment. The real-time analysis of multiple parameters together with the high sensitivity and high-throughput of our method thus enables new biological and therapeutic applications in the future.

Quantitative phase imaging through an ultra-thin lensless fiber endoscope.

Quantitative phase imaging (QPI) is a label-free technique providing both morphology and quantitative biophysical information in biomedicine. However, applying such a powerful technique to in vivo pathological diagnosis remains challenging. Multi-core fiber bundles (MCFs) enable ultra-thin probes for in vivo imaging, but current MCF imaging techniques are limited to amplitude imaging modalities. We demonstrate a computational lensless microendoscope that uses an ultra-thin bare MCF to perform quantitative phase imaging with microscale lateral resolution and nanoscale axial sensitivity of the optical path length. The incident complex light field at the measurement side is precisely reconstructed from the far-field speckle pattern at the detection side, enabling digital refocusing in a multi-layer sample without any mechanical movement. The accuracy of the quantitative phase reconstruction is validated by imaging the phase target and hydrogel beads through the MCF. With the proposed imaging modality, three-dimensional imaging of human cancer cells is achieved through the ultra-thin fiber endoscope, promising widespread clinical applications.

A customizable microfluidic platform for medium-throughput modeling of neuromuscular circuits.

Neuromuscular circuits (NMCs) are vital for voluntary movement, and effective models of NMCs are needed to understand the pathogenesis of, as well as to identify effective treatments for, multiple diseases, including Duchenne's muscular dystrophy and amyotrophic lateral sclerosis. Microfluidics are ideal for recapitulating the central and peripheral compartments of NMCs, but myotubes often detach before functional NMCs are formed. In addition, microfluidic systems are often limited to a single experimental unit, which significantly limits their application in disease modeling and drug discovery. Here, we developed a microfluidic platform (MFP) containing over 100 experimental units, making it suitable for medium-throughput applications. To overcome detachment, we incorporated a reactive polymer surface allowing customization of the environment to culture different cell types. Using this approach, we identified conditions that enable long-term co-culture of human motor neurons and myotubes differentiated from human induced pluripotent stem cells inside our MFP. Optogenetics demonstrated the formation of functional NMCs. Furthermore, we developed a novel application of the rabies tracing assay to efficiently identify NMCs in our MFP. Therefore, our MFP enables large-scale generation and quantification of functional NMCs for disease modeling and pharmacological drug targeting.