Exclusive Liquid Repellency: An Open Multi-Liquid-Phase Technology for Rare Cell Culture and Single-Cell Processing

Abstract

The concept of high liquid repellency in multi-liquid-phase systems (e.g., aqueous droplets in an oil background) has been applied to areas of biomedical research to realize intrinsic advantages not available in single-liquid-phase systems. Such advantages have included minimizing analyte loss, facile manipulation of single-cell samples, elimination of biofouling, and ease of use regarding loading and retrieving of the sample. In this paper, we present generalized design rules for predicting the wettability of solid-liquid-liquid systems (especially for discrimination between exclusive liquid repellency (ELR) and finite liquid repellency) to extend the applications of ELR. We then apply ELR to two model systems with open microfluidic design in cell biology: (1) in situ underoil culture and combinatorial coculture of mammalian cells in order to demonstrate directed single-cell multiencapsulation with minimal waste of samples as compared to stochastic cell seeding and (2) isolation of a pure population of circulating tumor cells, which is required for certain downstream analyses including sequencing and gene expression profiling.

Under oil open-channel microfluidics empowered by exclusive liquid repellency

“…The chambered coverglass was directly overlaid with oil. Silicone oil is the "right" oil to give the extreme wettability (i.e., ELR) based on our previous work (18). Tests on extraction of vital biomolecules (e.g., mRNA) from under oil and the quantitative polymerase chain reaction results in another previous work showed no obvious cargo loss during the processing (19).…”

“…Under the tested condition, the inlet microdroplet takes a torus shape to minimize the radius of curvature [R inlet = 2/ (1/R inlet-1 + 1/R inlet-2 ) = 0.67 mm, where R inlet-1 = h inlet , R inlet-2 = h inlet /2], and the outlet microdroplet is the shape of a spherical cap with a radius of curvature R outlet = 1.88 mm (defined by D outlet and h outlet ). P is defined by the Laplace pressure differential between the inlet and outlet spots (i.e., 2 oil/media /R inlet − 2 oil/media /R outlet , where  oil/media is the oil-media interfacial tension, about 41.8 mN/m at 25°C) (table S1) (18). The radius of curvature of the microchannel (R channel ) at t 0 (i.e., immediately after the microchannels filled with a solution by under oil sweep) can be calculated as R channel = 2/(1/R channel ⊥ + 1/R channel ∥ ) = 2 R channel ⊥ (table S2 and fig.…”

“…Recently, we developed under oil open microfluidics with the extreme wettability of exclusive liquid repellency (ELR) and double ELR (18)(19)(20). ELR is a phenomenon observed in solid-liquid-liquid three-phase systems, where a solid surface shows complete repellency to a liquid [with contact angle (CA) = 180°] when exposed to a second immiscible liquid.…”

Prostate cancer research: The next generation; report from the 2019 Coffey‐Holden Prostate Cancer Academy Meeting

“…Additional stacking with patient-matched M2 macrophages inhibited the migration of T cells, indicating the potential for functional assessment of a patient's tumor-microenvironment interaction (Lang et al, unpublished). [140][141][142] Pushing 3D culture systems to the next level, Lang et al have now begun to utilize exclusive and/or finite liquid repellency (ELR and FLR, respectively) [143][144][145] platforms to investigate tumor cell interactions with immune cells at the single-cell level. ELR/FLR enables the culture of single cells in microbubbles submerged in oil.…”

“…To date, proof-of-concept experiments using ELR have demonstrated successful single-cell culture of autologous human monocytes that have demonstrated differential potentials of individual monocytes to proliferate or not. 145 Further, the application of FLR allowed for successful single-cell culturing of human C4-2B prostate cancer cells, which demonstrated contrasting abilities of individual cells to proliferate, indicated by differential gene expression profiles. 144,145 ELR-based coculture experiments of human tumor-associated neutrophils with C4-2B tumoroids demonstrated heterogeneous interactions between tumoroid and neutrophil populations, including induction of tumor invasion versus apoptosis.…”