Fabrication of Microwell Arrays Based on Two-Dimensional Ordered Polystyrene Microspheres for High-Throughput Single-Cell Analysis

Abstract

This paper describes a method of fabricating rounded bottom microwell arrays (MA) in poly(dimethylsiloxane) (PDMS) by molding a monolayer of ordered polystyrene (PS) microspheres. PS microspheres were self-assembled on a glass slide and partially melted mainly from the bottom at 240 °C to increase adhesive force with the substrate. The partially melted PS arrays were used as master to generate MA. Microwell sizes are tunable in the 10-20 μm range with rounded bottoms; such a 3D structure is not readily obtainable through conventional soft lithography. Both adherent and nonadherent cell types can be retained in the microwells with high efficiency. As a demonstration of the advantage of real-time cell screening with this MA, single cell enzyme kinetic analysis was also carried out on trapped single cells. The PDMS MA may find applications in high-throughput drug screening, guided formation of cell clusters, and multicellular communication.

Hypoxia combined with spheroid culture improves cartilage specific function in chondrocytes

“…Different from the existing methods for fabricating concave microwell structure, such as ice-lithographic fabrication, 20 polymer micro-sphere 21 and elastic PDMS membrane deformation, 22 the approach we proposed is simply based on the SU-8 mold, which is quite simple, low cost and easy to operate, without the requirement of complicated instrumentation and procedures. In this work, we developed a straightforward strategy for rapid production of size controlled cell spheroids in a concave microwell array and resembled the in vivo-like physiological environment of cartilage by culturing chondrocytes under combined 3D spheroid and hypoxic conditions.…”

Microfluidic Devices in the Fast‐Growing Domain of Single‐Cell Analysis

“…They achieve the cell occupancy of ≈97 % along with the effective isolation of cells in the individual PEG micro‐well. Liu et al . have prepared 10–20 μm round bottom micro‐wells array in PDMS by molding a monolayer of ordered polystyrene (Figure a).…”

“… Microfluidic based single‐cell isolation (a) round‐bottom micro‐wells for single‐cell trapping using PS‐beads template, reprinted with permission from Ref. . Copyright© 2010 American Chemical Society.…”

“…They achieve the cell occupancyo f% 97 %a long with the effective isolation of cells in the individual PEG micro-well. Liu et al [87] have prepared 10-20 mmr ound bottom micro-wells array in PDMS by moldingamonolayer of ordered polystyrene (Figure 6a). They retained both adherent and non-adherent cells with high efficiency.C hen et al [88] have designed aptamer-encoded micro-wells to isolates ingle-tumor cells.…”

High-throughput Screening of Erratic Cell Volume Regulation Using a Hydrogel-based Single-cell Microwell Array

“…Polydimethyl siloxane (PDMS) microwell arrays were also prepared by casting PDMS prepolymer against a master that was made of a photoresist polymer 29 or selfassembled 2D polystyrene microbeads. 30 Yamamura et al used an injection molding to construct a polystyrene-based microwell array. 31 Among these previously reported methods, using a photocrosslinkable hydrogel array or a self-assembled microbead array is relatively simple and cost-effective to fabricate singlecell arrays.…”