Single cell trapping increasingly serves as a key manipulation technique in single cell analysis for many cutting-edge cell studies. Due to their inherent advantages, microfluidic devices have been widely used to enable single cell immobilization. To further improve the single cell trapping efficiency, this paper reports on a passive hydrodynamic microfluidic device based on the "least flow resistance path" principle with geometry optimized in line with corresponding cell types. Different from serpentine structure, the core trapping structure of the micro-device consists of a series of concatenated T and inverse T junction pairs which function as bypassing channels and trapping constrictions. This new device enhances the single cell trapping efficiency from three aspects: (1) there is no need to deploy very long or complicated channels to adjust flow resistance, thus saving space for each trapping unit; (2) the trapping works in a "deterministic" manner, thus saving a great deal of cell samples; and (3) the compact configuration allows shorter flowing path of cells in multiple channels, thus increasing the speed and throughput of cell trapping. The mathematical model of the design was proposed and optimization of associated key geometric parameters was conducted based on computational fluid dynamics (CFD) simulation. As a proof demonstration, two types of PDMS microfluidic devices were fabricated to trap HeLa and HEK-293T cells with relatively significant differences in cell sizes. Experimental results showed 100% cell trapping and 90% single cell trapping over 4 × 100 trap sites for these two cell types, respectively. The space saving is estimated to be 2-fold and the cell trapping speed enhancement to be 3-fold compared to previously reported devices. This device can be used for trapping various types of cells and expanded to trap cells in the order of tens of thousands on 1-cm(2) scale area, as a promising tool to pattern large-scale single cells on specific substrates and facilitate on-chip cellular assay at the single cell level.

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Optogenetics has been recently applied to manipulate the neural circuits of Caenorhabditis elegans (C. elegans) to investigate its mechanosensation and locomotive behavior, which is a fundamental topic in model biology. In most neuron-related research, free C. elegans moves on an open area such as agar surface. However, this simple environment is different from the soil, in which C. elegans naturally dwells. To bridge up the gap, this paper presents integration of optogenetic illumination of C. elegans neural circuits and muscular force measurement in a structured microfluidic chip mimicking the C. elegans soil habitat. The microfluidic chip is essentially a ∼1 × 1 cm2 elastomeric polydimethylsiloxane micro-pillar array, configured in either form of lattice (LC) or honeycomb (HC) to mimic the environment in which the worm dwells. The integrated system has four key modules for illumination pattern generation, pattern projection, automatic tracking of the worm, and force measurement. Specifically, two optical pathways co-exist in an inverted microscope, including built-in bright-field illumination for worm tracking and pattern generation, and added-in optogenetic illumination for pattern projection onto the worm body segment. The behavior of a freely moving worm in the chip under optogenetic manipulation can be recorded for off-line force measurements. Using wild-type N2 C. elegans, we demonstrated optical illumination of C. elegans neurons by projecting light onto its head/tail segment at 14 Hz refresh frequency. We also measured the force and observed three representative locomotion patterns of forward movement, reversal, and omega turn for LC and HC configurations. Being capable of stimulating or inhibiting worm neurons and simultaneously measuring the thrust force, this enabling platform would offer new insights into the correlation between neurons and locomotive behaviors of the nematode under a complex environment.

Leveraging microfluidics and nano-plasmonics, we present in this paper a new method employing a micro-nano-device that is capable of monitoring the dynamic cell-substrate attachment process at single cell level in real time without labeling. The micro-nano-device essentially has a gold thin film as the substrate perforated with periodic, near-cm2-area, template-stripped nano-holes, which generate plasmonic extraordinary optical transmission (EOT) with a high sensitivity to refractive index changes at the metal-dielectric interface. Using this device, we successfully demonstrated label-free and real-time monitoring of the dynamic cell attachment process for single mouse embryonic stem cell (C3H10) and human tumor cell (HeLa) by collecting EOT spectrum data during 3-hour on-chip culture. We further collected the EOT spectral shift data at the start and end points of measurement during 3-hour on-chip culture for 50 C3H10 and 50 HeLa cells, respectively. The experiment results show that the single cell attachment process of both HeLa and C3H10 cells follow the logistic retarded growth model, but with different kinetic parameters. Variations in spectral shift during the same culture period across single cells present new evidence for cell heterogeneity. The micro-nano-device provides a new, label-free, real-time, and sensitive, platform to investigate the cell adhesion kinetics at single cell level.

Single cell manipulation technology has been widely applied in biological fields, such as cell injection/enucleation, cell physiological measurement, and cell imaging. Recently, a biochip platform with a novel configuration of electrodes for cell 3D rotation has been successfully developed by generating rotating electric fields. However, the rotation platform still has two major shortcomings that need to be improved. The primary problem is that there is no on-chip module to facilitate the placement of a single cell into the rotation chamber, which causes very low efficiency in experiment to manually pipette single 10-micron-scale cells into rotation position. Secondly, the cell in the chamber may suffer from unstable rotation, which includes gravity-induced sinking down to the chamber bottom or electric-force-induced on-plane movement. To solve the two problems, in this paper we propose a new microfluidic chip with manipulation capabilities of single cell trap and single cell 3D stable rotation, both on one chip. The new microfluidic chip consists of two parts. The top capture part is based on the least flow resistance principle and is used to capture a single cell and to transport it to the rotation chamber. The bottom rotation part is based on dielectrophoresis (DEP) and is used to 3D rotate the single cell in the rotation chamber with enhanced stability. The two parts are aligned and bonded together to form closed channels for microfluidic handling. Using COMSOL simulation and preliminary experiments, we have verified, in principle, the concept of on-chip single cell traps and 3D stable rotation, and identified key parameters for chip structures, microfluidic handling, and electrode configurations. The work has laid a solid foundation for on-going chip fabrication and experiment validation.