High throughput microfluidics

Efficient tools for rapid antibiotic susceptibility testing (AST) are crucial for appropriate use of antibiotics, especially colistin, which is now often considered a last resort therapy with extremely drug resistant Gram-negative bacteria. Here, we developed a rapid, easy and miniaturized colistin susceptibility assay based on microfluidics, which allows for culture and high-throughput analysis of bacterial samples. Specifically, a simple microfluidic platform that can easily be operated was designed to encapsulate bacteria in nanoliter droplets and perform a fast and automated bacterial growth detection in 2 h, using standardized samples...

The incidence of urothelial carcinoma continues to rise annually, particularly among the elderly. Prompt diagnosis and treatment can significantly enhance patient survival and quality of life. Urine cytology remains a widely-used early screening method for urothelial carcinoma, but it still has limitations including sensitivity, labor-intensive procedures, and elevated cost. In recent developments, microfluidic chip technology offers an effective and efficient approach for clinical urine specimen analysis. Digital holographic microscopy, a form of quantitative phase imaging technology, captures extensive data on the refractive index and thickness of cells...

Tryptophan synthase catalyzes the synthesis of a wide array of noncanonical amino acids and is an attractive target for directed evolution. Droplet microfluidics offers an ultrahigh throughput approach to directed evolution (up to 107 experiments per day), enabling the search for biocatalysts in wider regions of sequence space with reagent consumption minimized to the picoliter volume (per library member). While the majority of screening campaigns in this format on record relied on an optically active reaction product, a new assay is needed for tryptophan synthase...

Immunoassays serve as powerful diagnostic tools for early disease screening, process monitoring, and precision treatment. However, the current methods are limited by high costs, prolonged processing times (>2 h), and operational complexities that hinder their widespread application in point-of-care testing. Here, we propose a novel centrifugo- p neumatic r eciprocating flowing coupled with spatial c onfinement strategy, termed PRCM , for ultrafast multiplexed immunoassay of pathogens on a centrifugal m icrofluidic platform...

Enumeration and phenotypic profiling of circulating tumor cells (CTCs) provide critical information for clinical diagnosis and treatment monitoring in cancer. To achieve this goal, an integrated system is needed to efficiently isolate CTCs from patient samples and sensitively evaluate their phenotypes. Such integration would comprise a high-throughput single-cell processing unit for the isolation and manipulation of CTCs and a sensitive and multiplexed quantitation unit to detect clinically relevant signals from these cells...

Cancer drug testing in animals is an extremely poor predictor of the drug's safety and efficacy observed in humans. Hence there is a pressing need for functional testing platforms that better predict traditional and immunotherapy responses in human, live tumor tissue or tissue constructs, and at the same time are compatible with the use of mouse tumor tissue to facilitate building more accurate disease models. Since many cancer drug actions rely on mechanisms that depend on the tumor microenvironment (TME), such platforms should also retain as much of the native TME as possible...

High-throughput single-cell analysis typically relies on the isolation of cells of interest in separate compartments for subsequent phenotypic or genotypic characterization. Using microfluidics, this is achieved by isolating individual cells in microdroplets or microwells. However, due to cell-to-cell variability in size, shape, and density, the cell capture efficiencies may vary significantly. This variability can negatively impact the measurements and introduce undesirable artifacts when trying to isolate and characterize heterogeneous cell populations...

Cell separation using microfluidics has become an effective method to isolate biological contaminants from bodily fluids and cell cultures, such as isolating bacteria contaminants from microalgae cultures and isolating bacteria contaminants from white blood cells. In this study, bacterial cells were used as a model contaminant in microalgae culture in a passive microfluidics device, which relies on hydrodynamic forces to demonstrate the separation of microalgae from bacteria contaminants in U and W-shaped cross-section spiral microchannel fabricated by defocusing CO2 laser ablation...

Several diseases affect the alveoli, and the efficacy of medical treatments and pharmaceutical therapies is hampered by the lack of pre-clinical models able to recreate in vitro the diseases. Microfluidic devices, mimicking the key structural and compositional features of the alveoli, offer several advantages to medium and high-throughput analysis of new candidate therapies. Here, we developed an alveolus-on-a-chip recapitulating the microanatomy of the physiological tissue by including the epithelium, the fibrous interstitial layer and the capillary endothelium...

Single-cell genomics has revolutionized tissue analysis by revealing the genetic program of individual cells. The key aspect of the technology is the use of barcoded beads to unambiguously tag sequences originating from a single cell. The generation of unique barcodes on beads is mainly achieved by split-pooling methods, which are labor-intensive due to repeated washing steps. Towards the automation of the split-pooling method, we developed a simple method to magnetize hydrogel beads. We show that these hydrogel beads provide increased yields and washing efficiencies for purification procedures...

Antibody drugs play a vital role in diagnostics and therapy. However, producing antibodies from mammalian cells is challenging owing to cellular heterogeneity, which can be addressed by applying droplet-based microfluidic platforms for high-throughput screening (HTS). Here, we designed an integrated system based on disulfide-bonded redox-responsive hydrogel beads (redox-HBs), which were prepared through enzymatic hydrogelation, to compartmentalize, screen, select, retrieve, and recover selected Chinese hamster ovary (CHO) cells secreting high levels of antibodies...

BACKGROUND: Small endosome-derived lipid nanovesicles (30-200 nm) are actively secreted by living cells and serve as pivotal biomarkers for early cancer diagnosis. However, the study of extracellular vesicles (EVs) requires isolation and purification from various body fluids. Although traditional EVs isolation and detection technologies are mature, they usually require large amount of sample, consumes long-time, and have relatively low-throughput. How to efficiently isolate, purify and detect these structurally specific EVs from body fluids with high-throughput remains a great challenge in in vitro diagnostics and clinical research...

Despite much progress, image processing remains a significant bottleneck for high-throughput analysis of microscopy data. One popular platform for single-cell time-lapse imaging is the mother machine, which enables long-term tracking of microbial cells under precisely controlled growth conditions. While several mother machine image analysis pipelines have been developed in the past several years, adoption by a non-expert audience remains a challenge. To fill this gap, we implemented our own software, MM3, as a plugin for the multidimensional image viewer napari...

Mounting evidence suggests that cell populations are extremely heterogeneous, with individual cells fulfilling different roles within the population. Flow cytometry (FC) is a high-throughput tool for single-cell analysis that works at high optical resolution. Sub-populations with unique properties can be screened, isolated and sorted through fluorescence-activated cell sorting (FACS), using intracellular fluorescent products or surface-tagged fluorescent products of interest. However, traditional FC and FACS methods cannot identify or isolate cells that secrete extracellular products of interest...

The arsM gene is a critical biomarker for the potential risk of arsenic exposure in paddy soil. However, on-site screening of arsM is limited by the lack of high-throughput point-of-use (POU) methods. Here, a multiplex CRISPR/Cas12a microfluidic paper-based analytical device (μPAD) was constructed for the high-throughput POU analysis of arsM , with cascade amplification driven by coupling crRNA-enhanced Cas12a and horseradish peroxidase (HRP)-modified probes. First, seven crRNAs were designed to recognize arsM , and their LODs and background signal intensities were evaluated...

BACKGROUND: Sperm selection, a key step in assisted reproductive technology (ART), has long been restrained at the preliminary physical level (morphology or motility); however, subsequent fertilization and embryogenesis are complicated biochemical processes. Such an enormous "gap" poses tough problems for couples dealing with infertility, especially patients with severe/total asthenozoospermia . METHODS: We developed a biochemical-level, automatic-screening/separation, smart droplet-TO-hydrogel chip (BLASTO-chip) for sperm selection...

Proteins as crucial components of cells are responsible for the majority of cellular processes. Sensitive and efficient protein detection enables a more accurate and comprehensive investigation of cellular phenotypes and life activities. Here, a protein sequencing method with high multiplexing, high throughput, high cell utilization, and integration based on digital microfluidics (DMF-Protein-seq) is proposed, which transforms protein information into DNA sequencing readout via DNA-tagged antibodies and labels single cells with unique cell barcodes...

Succinic acid (SA), one of the 12 top platform chemicals produced from biomass, is a precursor of various high value-added derivatives. Specially, 1 mol CO2 is assimilated in 1 mol SA biosynthetic route under anaerobic conditions, which helps to achieve carbon reduction goals. In this review, methods for enhanced CO2 fixation in SA production and utilization of waste biomass for SA production are reviewed. Bioelectrochemical and bioreactor coupling systems constructed with off-gas reutilization to capture CO2 more efficiently were highlighted...

Extensive research has demonstrated the potential of cell viscoelastic properties as intrinsic indicators of cell state, functionality, and disease. For this, several microfluidic techniques have been developed to measure cell viscoelasticity with high-throughput. However, current microchannel designs introduce complex stress distributions on cells, leading to inaccuracies in determining the stress-strain relationship and, consequently, the viscoelastic properties. Here, we introduce a novel approach using hyperbolic microchannels that enable precise measurements under a constant extensional stress and offer a straightforward stress-strain relationship, while operating at a measurement rate of up to 100 cells per second...

BACKGROUND: Single-cell droplet microfluidics is an important platform for high-throughput analyses and screening because it provides an independent and compartmentalized microenvironment for reaction or cultivation by coencapsulating individual cells with various molecules in monodisperse microdroplets. In combination with microbial biosensors, this technology becomes a potent tool for the screening of mutant strains. In this study, we demonstrated that a genetically engineered yeast strain that can fluorescently sense agonist ligands via the heterologous expression of a human G-protein-coupled receptor (GPCR) and concurrently secrete candidate peptides is highly compatible with single-cell droplet microfluidic technology for the high-throughput screening of new agonistically active peptides...