Single-Cell Point Constrictions for Reagent-Free High-Throughput Mechanical Lysis and Intact Nuclei Isolation.

Highly localized (point) constrictions featuring a round geometry with ultra-sharp edges in silicon have been demonstrated for the reagent-free continuous-flow rapid mechanical lysis of mammalian cells on a single-cell basis. Silicon point constrictions, robust structures formed by a single-step dry etching process, are arranged in a cascade along microfluidic channels and can effectively rupture cells delivered in a pressure-driven flow. The influence of the constriction size and count on the lysis performance is presented for fibroblasts in reference to total protein, DNA, and intact nuclei levels in the lysates evaluated by biochemical and fluoremetric assays and flow-cytometric analyses. Protein and DNA levels obtained from an eight-constriction treatment match or surpass those from a chemical method. More importantly, many intact nuclei are found in the lysates with a relatively high nuclei-isolation efficiency from a four-constriction treatment. Point constrictions and their role in rapid reagent-free disruption of the plasma membrane could have implications for integrated sample preparation in future lab-on-a-chip systems.

Railing cells along 3D microelectrode tracks for continuous-flow dielectrophoretic sorting.

We demonstrate a unique microfluidic device for continuous-flow cell sorting by railing target cells along physical tracks (electrode sidewalls) based on the combined effect of dielectrophoresis and hydrodynamic drag. The tracks are the raised digits of comb-like structures made of conducting bulk silicon as the electrodes. Unlike other volumetric electrodes, the structures feature a segmented sidewall profile with linear and concave segments forming the tracks and supporting columns, respectively. The interdigitated bulk electrodes lead to a built-in flow chamber in which the digits (tracks) extend downstream at a characteristic angle with respect to the flow, which runs through the passages between the columns. Target cells leaving the passages are levitated and docked against the tracks under positive dielectrophoresis and railed under hydrodynamic drag. Railing efficiency, as high as >95%, is reported against the activation voltage and flow rate for the designs 7°, 16°, and 26° as the track angles. A collection efficiency of about 86% is noted for both target (HCT116) and non-target cells (K562) in the 16° design at a sample flow rate of 8.3 µL min-1 and an activation voltage of 12.5 Vp at 200 kHz. This performance is comparable if not better than those obtained with thin-film surface microelectrodes and yet achieved here at an order of magnitude higher sample flow rate. This enhancement mainly arises from a considerably low drag along the tracks in relation to the chamber top or bottom surface where the thin-film electrodes would be typically placed.