Large-Area Electronics-Enabled High-Resolution Digital Microfluidics for Parallel Single-Cell Manipulation.

Large-area electronics as switching elements are an ideal option for electrode-array-based digital microfluidics. With support of highly scalable thin-film semiconductor technology, high-resolution digital droplets (diameter around 100 µm) containing single-cell samples can be manipulated freely on a two-dimensional plane with programmable addressing logic. In addition, single-cell generation and manipulation as foundations for single-cell research demand ease of operation, multifunctionality, and accurate tools. In this work, we reported an active-matrix digital microfluidic platform for single-cell generation and manipulation. The active device contained 26,368 electrodes that could be independently addressed to perform parallel and simultaneous droplet generation and achieved single-cell manipulation. We demonstrate a high-resolution digital droplet generation with a droplet volume limit of 500 pL and show the continuous and stable movement of droplet-contained cells for over 1 h. Furthermore, the success rate of single droplet formation was higher than 98%, generating tens of single cells within 10 s. In addition, a pristine single-cell generation rate of 29% was achieved without further selection procedures, and the droplets containing single cells could then be tested for on-chip cell culturing. After 20 h of culturing, about 12.5% of the single cells showed cell proliferation.

Enhanced absorbance detection system for online bacterial monitoring in digital microfluidics.

We report a fully integrated digital microfluidic absorbance detection system with an enhanced sensitivity for online bacterial monitoring. Through a 100 µm gap in the chip, our optical detection system has a detection sensitivity for a BCA protein concentration of 0.1 mg mL-1. The absorbance detection limit of our system is 1.4 × 10-3 OD units, which is one order of magnitude better than that of the existing studies. The system's linear region is 0.1-7 mg mL-1, and the dynamic range is 0-25 mg mL-1. We measured the growth curves of wild-type and E. coli transformed with resistance plasmids and mixed at different ratios on chip. We sorted out the bacterial species including highly viable single cells based on the difference in absorbance data of growth curves. We explored the changes in the growth curves of E. coli under different concentrations of resistant media. In addition, we successfully screened for the optimal growth environment of the bacteria, in which the growth rate of PET30a-DH5α (in a medium with 33 µg mL-1 kanamycin resistance) was significantly higher than that of a 1 mg mL-1 resistance medium. In conclusion, the enhanced digital microfluidic absorbance detection system exhibits exceptional sensitivity, enabling precise bacterial monitoring and growth curve analysis, while also laying the foundation for DMF-based automated bioresearch platforms, thus advancing research in the life sciences.

Open Thermal Control System for Stable Polymerase Chain Reaction on a Digital Microfluidic Chip.

In this paper, a digital microfluidic thermal control system was introduced for the stable polymerase chain reaction (PCR). The system consists of a thermoelectric cooler unit, a thermal control board, and graphical-user-interface software capable of simultaneously achieving temperature control and on-chip droplet observation. A fuzzy proportional-integral-derivative (PID) method was developed for this system. The simulation analysis was performed to evaluate the temperature of different reagents within the chip. Based on the results, applying fuzzy PID control for PCR will enhance the thermal stability by 67.8% and save the time by 1195 s, demonstrating excellent dynamic response capability and thermal robustness. The experimental results are consistent with the simulation results on the planar temperature distribution, with a data consistency rate of over 99%. The PCR validation was carried out on this system, successfully amplifying the rat GAPDH gene at a concentration of 193 copies/µL. This work has the potential to be useful in numerous existing lab-on-a-chip applications.

AIEgens-enhanced rapid sensitive immunofluorescent assay for SARS-CoV-2 with digital microfluidics.

BACKGROUND: Sensitive and rapid antigen detection is critical for the diagnosis and treatment of infectious diseases, but conventional ELISAs including chemiluminescence-based assays are limited in sensitivity and require many operation steps. Fluorescence immunoassays are fast and convenient but often show limited sensitivity and dynamic range. RESULTS: To address the need, an aggregation-induced emission fluorgens (AIEgens) enhanced immunofluorescent assay with beads-based quantification on the digital microfluidic (DMF) platform was developed. Portable DMF devices and chips with small electrodes were fabricated, capable of manipulating droplets within 100 nL and boosting the reaction efficiency. AIEgen nanoparticles (NPs) with high fluorescence and photostability were synthesized to enhance the test sensitivity and detection range. The integration of AIEgen probes, transparent DMF chip design, and the large magnetic beads (10 µm) as capture agents enabled rapid and direct image-taking and signal calculation of the test result. The performance of this platform was demonstrated by point-of-care quantification of SARS-CoV-2 nucleocapsid (N) protein. Within 25 min, a limit of detection of 5.08 pg mL-1 and a limit of quantification of 8.91 pg mL-1 can be achieved using <1 µL sample. The system showed high reproducibility across the wide dynamic range (10-105 pg mL-1), with the coefficient of variance ranging from 2.6% to 9.8%. SIGNIFICANCE: This rapid, sensitive AIEgens-enhanced immunofluorescent assay on the DMF platform showed simplified reaction steps and improved performance, providing insight into the small-volume point-of-care testing of different biomarkers in research and clinical applications.

Active-matrix digital microfluidics design for field programmable high-throughput digitalized liquid handling.

Digital liquid sample handling is an enabling tool for cutting-edge life-sciences research. We present here an active-matrix thin-film transistor (TFT) based digital microfluidics system, referred to as Field Programmable Droplet Array (FPDA). The system contains 256 × 256 pixels in an active area of 10.65 cm2, which can manipulate thousands of addressable liquid droplets simultaneously. By leveraging a novel TFT device and circuits design solution, we manage to programmatically manipulate droplets at single-pixel level. The minimum achievable droplet volume is around 0.5 nL, which is two orders of magnitude smaller than the smallest droplet ever reported on active-matrix digital microfluidics. The movement of droplets can be either pre-programmed or controlled in real-time. The FPDA system shows great potential of the ubiquitous thin-film electronics technology in digital liquid handling. These efforts will make it possible to create a true programmable lab-on-a-chip device to enable great advances in life science research.

Advanced design and applications of digital microfluidics in biomedical fields: An update of recent progress.

Significant breakthroughs have been made in digital microfluidic (DMF)-based technologies over the past decades. DMF technology has attracted great interest in bioassays depending on automatic microscale liquid manipulations and complicated multi-step processing. In this review, the recent advances of DMF platforms in the biomedical field were summarized, focusing on the integrated design and applications of the DMF system. Firstly, the electrowetting-on-dielectric principle, fabrication of DMF chips, and commercialization of the DMF system were elaborated. Then, the updated droplets and magnetic beads manipulation strategies with DMF were explored. DMF-based biomedical applications were comprehensively discussed, including automated sample preparation strategies, immunoassays, molecular diagnosis, blood processing/testing, and microbe analysis. Emerging applications such as enzyme activity assessment and DNA storage were also explored. The performance of each bioassay was compared and discussed, providing insight into the novel design and applications of the DMF technology. Finally, the advantages, challenges, and future trends of DMF systems were systematically summarized, demonstrating new perspectives on the extensive applications of DMF in basic research and commercialization.

"One-to-three" droplet generation in digital microfluidics for parallel chemiluminescence immunoassays.

In digital microfluidics, droplet generation is a fundamental operation for quantitative liquid manipulation. The generation of well-defined micro-droplets on a chip with restricted device geometries has become a real obstacle for digital microfluidics platforms to be used in parallel for in vitro diagnostic applications. Here, we propose a "one-to-three" droplet splitting technique that is able to generate sub-microlitre droplets beyond the "well-known" geometry limit in electrowetting-on-dielectric digital microfluidics. Accordingly, we realized an on-chip magnetic bead chemiluminescence immunoassay for parallel detection with the "one-to-three" technique. With the help of the generated micro droplets, we were able to retain the magnetic beads by a significantly reduced magnetic force. We have shown the detection of five B-type natriuretic peptide analyte samples on a single chip for around 10 minutes. The correlation coefficient of the calibration curve was 0.9942, and the detection limit was lower than 5 pg mL-1.

Trends and recent development of the microelectrode arrays (MEAs).

In this paper, we reviewed the history of microelectrode arrays (MEAs), compared different microfabrication techniques applied to modern MEAs in terms of their material characters, device properties and application scenarios. Then we discussed the biocompatibility of different MEAs as well as corresponding strategy of improvement. At last, we analyzed the growing trend of MEAs' technical route, expected application of MEAs in the field of Electrical impedance tomography (EIT).