A Facile and Rapid Route to Self-Digitization of Samples into a High Density Microwell Array for Digital Bioassays

Abstract

Digital bioassays are powerful methods to detect rare analytes from complex mixtures and study the temporal processes of individual entities within biological systems. In digital bioassays, a crucial first step is the discretization of samples into a large number of identical independent partitions. Here, we developed a rapid and facile sample partitioning method for versatile digital bioassays. This method is based on a detachable self-digitization (DSD) chip which couples a reversible assembly configuration and a predegassing-based self-pumping mechanism to achieve an easy, fast and large-scale sample partitioning. The DSD chip consists of a channel layer used for loading sample and a microwell layer used for holding the sample partitions. Benefitting from its detachability, the chip avoids a lengthy oil flushing process used to remove the excess sample in loading channels and can compartmentalize a sample into more than 100,000 wells of picoliter volume with densities up to 14,000 wells/cm2 in less than 30 s. We also demonstrated the utility of the proposed method by applying it to digital PCR and digital microbial assays.

Facile and Low-Waste Self-Digitization of Samples by Oil-Triggered, Template-Confined Dewetting on a Networked Microwell Array

“…First, the inlet of the chip was sealed with tape, and then the sealed chip was placed in a vacuum condition (~ 10 kPa) for 1 h to build up a negative pressure in the closed microchannel/ microwell system for self-priming. 27 After removal of the chip from the vacuum, a pipette tip holding aqueous sample (~35 μL) was immediately inserted through the tape and into the inlet of the chip. Driven by the negative pressure created in the chip during the degassing process, the sample solution was automatically sucked into the microchannels and quickly filled all the available space in the closed system including microchannels and microwells.…”

“…Driven by the negative pressure created in the chip during the degassing process, the sample solution was automatically sucked into the microchannels and quickly filled all the available space in the closed system including microchannels and microwells. [23][24][25][26][27][28][29] Subsequently, a few drops of oil having high wettability to PDMS (e.g., silicone oil or hexadecane) were added to the oil reservoir. Once the sample filled all of the channels and microwells, the cover layer was peeled off from the microstructured layer.…”

“…More recently, in order to lessen reliance on complex external pumping equipment and simplify the fluid operation for end-users, a degassing-driven self-pumping mechanism was recently introduced for "static" sample digitization. [23][24][25][26][27] By employing the air absorption of the pre-degassed polydimethylsiloxane (PDMS) substrate to create pumping pressure, this power-free self-digitization technology has minimized the number of liquid control components, which is favorable to portable applications. In most cases, the power-free self-digitization devices consist of an array of microchambers, each row of microchambers connecting to a main microchannel.…”

“…Therefore, it remains challenging for microfluidic platforms to achieve rapid, cost-effective, and low-sample-loss digitization. To address these challenges, we recently presented a rapid and facile method for spontaneous sample digitization, 27 which uses a reversible channel-well assembly chip coupled with predegassing-based self-pumping mechanism to achieve an easy, fast, and large-scale sample partitioning. However, due to the detachable channel-well configuration, it requires tedious alignment and assembly which can hinder non-specialist users from using this sample digitization method.…”

Static Quasi-Double Emulsions as Programmable Microcrystallizers for Controllable Polymorphic Crystallization

“…To generate highly reproducible, uniform, and stable double emulsion structures for facilely and flexibly controlling the polymorphic crystallization, we develop a QDE chip based on our previous works. 28 The QDE chip is schematically shown in Figure 1b, which is reversibly assembled from three layers: a polydimethylsiloxane (PDMS)based channel layer, a PDMS-based well layer, and a supporting glass substrate. The channel layer contains an array of 667 parallel microfluidic channels used for IAP delivery, and all channels connect to an inlet port via a main channel.…”