Microfluidic Chip Research Articles

Combinatorial sample preparation platform for droplet-based applications in microbiology

Droplet microfluidics has demonstrated immense potential in microbiological studies due to its unique features, such as miniaturization, compartmentalization, and parallelization. Multiplexing droplet content allows the investigation of various experimental conditions in a highly parallelized manner. Yet, droplet library generation and tracking remain challenging in high-throughput screening. The introduction of distinct reagents into droplets necessitates precise control over droplet flow in a microfluidic chip, limiting the throughput to a few reagents. Additionally, tracking individual droplets is complex due to their fast dynamics. To address these challenges, we have developed a multiplexing platform for automated sample preparation, enabling on-demand merging and mixing of reagents for fine-tuning the sample compositions for droplet generation. A coding space with 169 optical barcodes can be realized by the pairwise combination of four fluorescence dyes at six concentration levels to encode droplet populations as required by the experimental design. A machine-learning algorithm has been employed to identify distinct droplet populations. As proof of concept, we conducted an antibiotic susceptibility assay on an E. coli strain to screen for susceptibility of four antibiotics and determine minimum inhibitory concentrations in one experiment. Utilizing the on-demand sample preparation, optical barcodes, and machine-learning analysis, our setup provides a rapid, straightforward, and reliable multiplexing capability for numerous microbial and biochemical applications.

The impact of choledochal cysts on bile fluid dynamics: A perspective using computational fluid dynamics and surface mapping technique

Choledochal cysts (CCs) are an important risk factor for cholangiocarcinoma, though their etiology remains debated. Given the vital role of bile fluid in digestive processes within the biliary system, examining such mechanisms from the perspective of bile fluid dynamics may offer additional insights for clinical use. This study utilized magnetic resonance imaging (MRI)-based patient-specific scans for detailed reconstruction and further employed the computational fluid dynamic method to assess the physiological functions of each system, including refilling and emptying processes. The impact of bile rheological property was also examined. Key biomechanical parameters—pressure and wall shear stress (WSS)—were displayed on a two-dimensional plane via surface mapping for enhanced visualization and comparative analysis. Outcomes demonstrated a significant reduction in bile flow velocity in CCs patients due to common bile duct's anatomical features and bile's shear-thinning, non-Newtonian nature, with a notable increase in pressure drop observed. In healthy biliary systems, WSS variations were minimal; however, in CCs patients, extreme WSS differences were found, with the highest WSS in the segmental bile duct and the lowest in the dilatation area, presenting a magnitude difference of approximately 1000. CCs one showed WSS levels 100–250 times higher than healthy ones in the common bile duct. Bile rheological properties substantially affect pressure and WSS patterns, particularly WSS, where pathological bile caused a tenfold increase in WSS compared to healthy bile. The findings aimed to enhance the understanding of biliary fluid mechanics in CCs and offer insights into selected fluidic variables for future microfluidic chip experiments.

A portable microfluidic electrochemical sensor with nonlinear fit strategy for wide-range uric acid detection

In this work, we have developed a portable electrochemical sensing device controlled by a smartphone through Bluetooth, which was applied for regular uric acid (UA) monitoring in prevention and healthcare management. On the core lies a nonlinear fitting model (I = k·(N − exp(−K·cbulk + b))) which is proposed for the first time and given from the differential equation based on the theoretical analysis. We believe this model can reflect the intrinsic relationship between the obtained current and the bulk analyte concentration, therefore leading to an expanding UA detection range (10-fold) compared to the widely used linear standard curve model and reducing the requirements for the modification of the electrode materials. Besides, the integration of the microfluidic chip promoted the removal of UA oxidation products at the electrode surface, yielding excellent sensing stability with a relative standard deviation (RSD) lower than 1 % during 10 consecutive runs. Consequently, we acquired the relationship between Square Wave Voltammetry (SWV) current and UA concentration (I = 15.27*(2.05 − exp(−0.00108*(cbulk − 663.5))), R2 = 0.9999) in the range of 5 to 1000 μM with a limit of detection (LoD) of 2.4 μM using a completely unmodified screen-printed carbon electrode (SPCE). Proof of concept experiments using 25× diluted human urine spiked with UA yielded a recovery rate of 87.5–101.4 % and a satisfactory selectivity result, which is within the value expected for clinical use, indicating the potential of the developed instrument and nonlinear fitting model for urinary UA detection.

3D printed microfluidic chip integrated with nanointerferometer for multiplex detection of foodborne pathogens

The current foodborne pathogen detection methods, such as culture-based methods, polymerase chain reaction, and optical and electrochemical biosensors with nucleic acid, have high sensitivity and selectivity. However, they are slow, expensive, and require well-trained operators. In this study, we utilized a 3D printer to develop a novel chip with an aptamer-based nanointerferometer capable of identifying four distinct foodborne pathogens: Listeria monocytogenes, Escherichia coli, Salmonella typhimurium, and Staphylococcus aureus. The aptamer sensor on the chip achieved a limit-of-detection of 10 colony forming unit (CFU)/ml. With its high sensitivity and specificity, this chip offers a cost-effective platform for distinguishing and screening different foodborne pathogens.

Enabling accessibility to the CellScape spatial biology workflow for researchers using standard histology protocols and pre-slided specimens.

e15185 Background: Solid tumor specimen collection during clinical trials often follows a standard histology protocol, ending in the generation of FFPE sections mounted onto 1 mm histology slides. Though a common sample format for researchers developing tissue-based biomarkers, slides present a data quality challenge for multiplexed spatial biology workflows. Here we present data describing an innovative flow cell design fully compatible with pre-slided tissue specimens. We collected spatial biology data using this slide-compatible flow cell and compared it to the gold standard direct mounting on coverglass. Several rounds of marker staining and imaging demonstrate equivalency in method outcomes including image resolution, signal to noise, and re-interrogation capacity. Methods: FFPE biopsy tissue specimens (tumor punch biopsies and a tonsil section) were mounted on standard histology slides. These specimens were incorporated into flow cells using a new glass coverslip and porting device, CellScape™ Slide, that pairs with a standard histology slide to form a microfluidic chamber. After mounting, each sample was probed for consensus CD markers (CD45, CD3, CD4,CD8,CD20, etc.) through several rounds of iterative immunofluorescence staining using CellScape™ Precise Spatial Multiplexing. The resulting multi-layered whole slide image dataset was then interrogated for spatial resolution and signal-to-noise using digital image analysis. Results: Spatial resolution and signal-to-noise ratio (SNR) were assessed through comparison to data collected from similar samples using the existing CellScape microfluidic chip. The lowest intensity signal collected with at least a 30:1 SNR (acceptable for image analysis) was equivalent or lower in the CellScape Slide preparation (equivalent sensitivity). The highest intensity signal collected was equivalent between the two flow cell designs. Comparing stain quality (specificity, sensitivity) among markers stained through multiple rounds of iterative staining, imaging, and signal remove via photoinactivation showed that the CellScape Slide flow cell is on par with or better than the existing CellScape chip. Further, the usable clear window of the CellScape Slide flow chamber, measuring 16 mm x 44 mm, was found to be greater than twice that of the existing chip. Literature review determined this to also be the largest imaging window available among commercially available spatial biology platforms. Conclusions: Assessment of the CellScape Slide flow cell, innovative in providing access to the CellScape spatial biology workflow for pre-mounted FFPE tissue, determined that the optical performance of the new flow cell is at least on par with the existing flow cell. Further, the usable window is over twice the area, and the new flow cell is fully capable of supporting re-interrogation of the sample.

Droplet array with microfluidic concentration gradient (DA-MCG) for 2-dimensional reaction condition screening

Droplet microfluidic technology can use each microdroplet as a microreactor, which has the advantages of low reagent dosage, less cross contamination, and fast reaction time. Combining concentration gradient generation with droplet formation and capture to form a two-dimensional reaction condition screening platform (including reactant concentration and reaction time) is expected to broaden the application range of microfluidic screening. In this work, a microfluidic chip that can dynamically generate and capture microdroplets and form static microdroplet array was designed and fabricated. An optimized Christmas tree structure by adjusting the horizontal channel length ratio was used to generate a chemical concentration gradient while obtaining a uniform outlet flow rate, forming a droplet array with different concentrations. The performance of droplet array with microfluidic concentration gradient (DA-MCG) was verified using sodium fluorescein as a model reagent. The chromogenic reaction of NaOH and phenolphthalein, and luminol chemiluminescence reaction were used to verify the two-dimensional screening ability of DA-MCG. The results indicated that the DA-MCG has the potential to be applied in the field of multi-dimensional drug screening.

In vitro assessment of ferroptosis of cells exposed to cigarette smoke aerosol using a self-designed on-chip evaluation system based on gas–liquid dual-dimensional exposure

Aerosol pollutants significantly cause health concerns. Herein, we established an original real-time aerosol exposure system that used a self-designed bionic-lung microfluidic chip. The chip features a 4 × 4 intersecting array within gas and liquid layers, creating 16 distinct microenvironments. A membrane situated between the layers offers attachment for cells and establishes a gas–liquid interface. This design provides a reliable screening capacity for investigating the biological effects of aerosol exposure in vitro by manipulating the gas and/or liquid conditions. Using this system, we validated that cigarette smoke (CS) aerosol triggered a concentration- and time-dependent reduction in cell viability and intracellular glutathione levels, accompanied by an increase in intracellular reactive oxygen species and Fe2+. Furthermore, CS aerosol significantly downregulated the expression of GPX4, SLC7A11, and FTL mRNA while inducing a notable increase in that of ACSL4 mRNA. Additionally, CS aerosol markedly stimulated the release of proinflammatory cytokines. Crucially, the ferroptosis inhibitor deferoxamine mesylate reversed these biological indicators. These results demonstrate that our novel bionic-lung chip presents a suitably achievable approach to investigate the biological effects induced by aerosol exposure.

A facile single-cell patterning strategy based on harbor-like microwell microfluidics

Single-cell analysis is an effective method for conducting comprehensive heterogeneity studies ranging from cell phenotype to gene expression. The ability to arrange different cells in a predetermined pattern at single-cell resolution has a wide range of applications in cell-based analysis and plays an important role in facilitating interdisciplinary research by researchers in various fields. Most existing microfluidic microwell chips is a simple and straightforward method, which typically use small-sized microwells to accommodate single cells. However, this method imposes certain limitations on cells of various sizes, and the single-cell capture efficiency is relatively low without the assistance of external forces. Moreover, the microwells limit the spatiotemporal resolution of reagent replacement, as well as cell-to-cell communication. In this study, we propose a new strategy to prepare a single-cell array on a planar microchannel based on microfluidic flip microwells chip platform with large apertures (50 μm), shallow channels (50 μm), and deep microwells (50 μm). The combination of three configuration characteristics contributes to multi-cell trapping and a single-cell array within microwells, while the subsequent chip flipping accomplishes the transfer of the single-cell array to the opposite planar microchannel for cells adherence and growth. Further assisted by protein coating of bovine serum albumin and fibronectin on different layers, the single-cell capture efficiency in microwells is achieved at 92.1% ± 1%, while ultimately 85% ± 3.4% on planar microchannel. To verify the microfluidic flip microwells chip platform, the real-time and heterogeneous study of calcium release and apoptosis behaviours of single cells is carried out. To our knowledge, this is the first time that high-efficiency single-cell acquisition has been accomplished using a circular-well chip design that combines shallow channel, large aperture and deep microwell together. The chip is effective in avoiding the shearing force of high flow rates on cells, and the large apertures better allows cells to sedimentation. Therefore, this strategy owns the advantages of easy preparation and user-friendliness, which is especially valuable for researchers from different fields.

A High-Throughput Circular Tumor Cell Sorting Chip with Trapezoidal Cross Section.

Circulating tumor cells are typically found in the peripheral blood of patients, offering a crucial pathway for the early diagnosis and prediction of cancer. Traditional methods for early cancer diagnosis are inefficient and inaccurate, making it difficult to isolate tumor cells from a large number of cells. In this paper, a new spiral microfluidic chip with asymmetric cross-section is proposed for rapid, high-throughput, label-free enrichment of CTCs in peripheral blood. A mold of the desired flow channel structure was prepared and inverted to make a trapezoidal cross-section using a micro-nanotechnology process of 3D printing. After a systematic study of how flow rate, channel width, and particle concentration affect the performance of the device, we utilized the device to simulate cell sorting of 6 μm, 15 μm, and 25 μm PS (Polystyrene) particles, and the separation efficiency and separation purity of 25 μm PS particles reached 98.3% and 96.4%. On this basis, we realize the enrichment of a large number of CTCs in diluted whole blood (5 mL). The results show that the separation efficiency of A549 was 88.9% and the separation purity was 96.4% at a high throughput of 1400 μL/min. In conclusion, we believe that the developed method is relevant for efficient recovery from whole blood and beneficial for future automated clinical analysis.

Study on Circulating Tumor Cell Detection System Based on Microfluidic Chip

To achieve high throughput and high detection rate of circulating tumor cells (CTCs) in human peripheral blood, and to provide efficient and accurate early screening for cancer patients. A microfluidic chip with the integration of sorting, enrichment and detection was designed, and CTCs at the single cell level were detected by fluorescence detection system to obtain the number of CTCs in samples. The peripheral blood samples after lysed red blood cells were used for 6 experiments. When the injection rate reached 0.2 mL/h, CTCs could reach the best detection rate of 78.6%, and the correlation coefficient within the group was above 0.8. CTCs detection system can achieve high detection rate and has good reliability, which can provide a reliable reference for clinical research in related fields.

Rapid Microfluidic Immuno-Biosensor Detection System for the Point-of-Care Determination of High-Sensitivity Urinary C-Reactive Protein.

A microfluidic immuno-biosensor detection system consisting of a microfluidic spectrum chip and a micro-spectrometer detection device is presented for the rapid point-of-care (POC) detection and quantification of high-sensitivity C-reactive protein (hs-CRP) in urine. The detection process utilizes a highly specific enzyme-linked immunosorbent assay (ELISA) method, in which capture antibodies and detection antibodies are pre-deposited on the substrate of the microchip and used to form an immune complex with the target antigen. Horseradish peroxidase (HRP) is added as a marker enzyme, followed by a colorimetric reaction using 3,3',5,5'-tetramethylbenzidine (TMB). The absorbance values (a.u.) of the colorimetric reaction compounds are measured using a micro-spectrometer device and used to measure the corresponding hs-CRP concentration according to the pre-established calibration curve. It is shown that the hs-CRP concentration can be determined within 50 min. In addition, the system achieves recovery rates of 93.8-106.2% in blind water samples and 94.5-104.6% in artificial urine. The results showed that the CRP detection results of 41 urine samples from patients with chronic kidney disease (CKD) were highly consistent with the conventional homogeneous particle-enhanced turbidimetric immunoassay (PETIA) method's detection results (R2 = 0.9910). The experimental results showed its applicability in the detection of CRP in both urine and serum. Overall, the results indicate that the current microfluidic ELISA detection system provides an accurate and reliable method for monitoring the hs-CRP concentration in point-of-care applications.

Mechanical Features of PMMA Microfluidic Chips Implemented by Carbon Dioxide Laser

Mechanical Features of PMMA Microfluidic Chips Implemented by Carbon Dioxide Laser

A versatile dilution-treatment-detection microfluidic chip platform for rapid In vitro lung cancer drug combination sensitivity evaluation

Combination drug therapy represents an effective strategy for treating certain drug-resistant and intractable cancer cases. However, determining the optimal combination of drugs and dosages is challenging due to clonal diversity in patients' tumors and the lack of rapid drug sensitivity evaluation methods. Microfluidic technology offers promising solutions to this issue. In this study, we propose a versatile microfluidic chip platform capable of integrating all processes, including dilution, treatment, and detection, for in vitro drug sensitivity assays. This platform innovatively incorporates several modules, including automated discrete drug logarithmic concentration generation, on-chip cell perfusion culture, and parallel drug treatments of cancer cell models. Moreover, it is compatible with microplate readers or high-content imaging systems for swift detection and automated monitoring, simplifying on-chip drug evaluation. Proof of concept is demonstrated by assessing the in vitro potency of two drugs, cisplatin, and etoposide, against the lung adenocarcinoma A549 cell line, under both single-drug and combination treatment conditions. The findings reveal that, compared to conventional microplate approaches with static cultivation, this on-chip automated perfusion bioassays yield comparable IC50 values with lower variation and a 50 % reduction in drug preparation time. This versatile dilution-treatment-detection microfluidic platform offers a promising tool for rapid and precise drug assessments, facilitating in vitro drug sensitivity evaluation in personalized cancer chemotherapy.

Target Tracking Two Degrees of Freedom State Feedback Control for Continuous Flow Microfluidic Chips Temperature Controller

Microfluidic chips represent a cutting-edge technology for manipulating fluids within micrometer-scale spaces and are gradually becoming a new favorite platform in life science research. Precise and fast zonal temperature control is essential for accelerating biological experiments. However, current multi-channel temperature controllers typically rely on multiple channel sets to achieve single set-point control, which results in discrepancies between the fluid temperature distribution and sensor temperature due to the distributed temperature field in the fluid channel. To estimate the actual temperature and implement gradient temperature control, this paper introduces an extension of the target tracking (TT) two degrees of freedom (2DOF) state feedback control (SFC) method, followed by a presentation of simulation and experimental results. Through comparisons with an enhanced PID system in both simulation and experimentation, the paper demonstrates an 8.96% reduction in the maximum temperature difference across different regions and a 27.89% decrease in the time taken to reach various temperatures. This solution effectively addresses the existing challenges in temperature control for microfluidic chips, offering a more precise and stable control within the desired temperature range.

Design, Fabrication and Testing of A Novel Thermally-Actuated Tesla Valve (TATV): A Hybrid Microvalve

Abstract A novel thermally-actuated hybrid microvalve (similar to a Tesla Valve configuration) was designed, fabricated, assembled, and tested using soft lithography-based approaches. The modified design integrates the "normally open and closed" hybrid configurations of Tesla Valves with a thermo-pneumatic (or thermal-hydraulic) actuator microfabricated in-situ that modulates the diodicity of the microvalve apparatus in the microfluidic chip. Diodicity (Di) is defined as the ratio of flow rate in the forward direction to that of the reverse direction (for a constant value of pressure drop that is imposed on a microvalve device). The results from the study successfully demonstrated the operation of an array of Tesla Valves that are normally open in forward direction and marginally closed in reverse direction at room temperature (i.e., with Di > 1, the flow resistance values were different when the inlet and outlet ports were swapped). When the microfluidic chip was heated (at steady-state conditions with a nominal temperature of ~30°C), the diodicity virtually vanished (i.e., Di ≈ 1) resulting in both reverse and forward directions being normally open (or having the same flow resistance - irrespective of the flow direction).

Lab on a chip (LOC) platform for drug screening against the intracellular forms of Trypanosoma cruzi

Trypanosoma cruzi is the etiological agent of Chagas disease, endemic in Latin America with recent spreading to non-endemic areas due to human migration. The low efficacy and substantial prevalence of side effects of the approved medications benznidazole (BZN) and nifurtimox observed mainly during the chronic phase of Chagas disease, when most patients receive their diagnosis, demonstrate the need to find new therapeutic compounds to address this neglected global health disease. Here, we developed a platform for screening the antitrypanosomal activity of compounds over intracellular amastigotes using microfluidics, LOC devices, and advanced image analysis algorithms. This platform allows the simultaneous testing of six different concentrations of a compound and the automatic and high-precision processing of the images obtained during the experimental phase, providing quickly and easily a multitude of statistical data for the screening of new drugs. To assess the developed platform, we analyzed the effects of BZN as a reference drug and compared the dose–response curves with those obtained with a flow cytometry-based method, obtaining similar IC50 results. Furthermore, a collection of compounds obtained by chemical synthesis from a combinatorial library was tested over intracellular amastigotes in infected Vero cells.

Evaluation of the QMAC-dRAST System Version 2.5 for Rapid Antimicrobial Susceptibility Testing of Gram-Negative Bacteria From Positive Blood Culture Broth and Subcultured Colony Isolates.

Rapid antimicrobial susceptibility testing (AST) for bloodstream infections (BSIs) facilitates the optimization of antimicrobial therapy, preventing antimicrobial resistance and improving patient outcomes. QMAC-dRAST (QuantaMatrix Inc., Korea) is a rapid AST platform based on microfluidic chip technology that performs AST directly using positive blood culture broth (PBCB). This study evaluated the performance of QMAC-dRAST for Gram-negative bacteria using PBCB and subcultured colony isolates, comparing it with that of VITEK 2 (bioMérieux, France) using broth microdilution (BMD) as the reference method. We included 141 Gram-negative blood culture isolates from patients with BSI and 12 carbapenemase-producing clinical isolates of Enterobacterales spiked into blood culture bottles. QMAC-dRAST performance was evaluated using PBCB and colony isolates, whereas VITEK 2 and BMD were tested only on colony isolates. For PBCB, QMAC-dRAST achieved 92.1% categorical agreement (CA), 95.3% essential agreement (EA), with 1.8% very major errors (VMEs), 3.5% major errors (MEs), and 5.2% minor errors (mEs). With colony isolates, it exhibited 92.5% CA and 95.1% EA, with 2.0% VMEs, 3.2% MEs, and 4.8% mEs. VITEK 2 showed 94.1% CA and 96.0% EA, with 4.3% VMEs, 0.4% MEs, and 4.3% mEs. QMAC-dRAST yielded elevated error rates for specific antimicrobial agents, with high VMEs for carbapenems and aminoglycosides. The median time to result for QMAC-dRAST was 5.9 h for PBCB samples and 6.1 h for subcultured colony isolates. The QMAC-dRAST system demonstrated considerable strengths and comparable performance to the VITEK 2 system; however, challenges were discerned with specific antimicrobial agents, underlining a necessity for improvement.

A bibliometric analysis of 3D microfluidic chip in cancer.

A bibliometric analysis of 3D microfluidic chip in cancer.

An ultrahigh frequency dielectric sensor for microdroplet detection using a split ring resonator-based microfluidic chip

An ultrahigh frequency dielectric sensor for microdroplet detection using a split ring resonator-based microfluidic chip

Progress and application of lung-on-a-chip for lung cancer

Lung cancer is a malignant tumour with the highest incidence and mortality worldwide. Clinically effective therapy strategies are underutilized owing to the lack of efficient models for evaluating drug response. One of the main reasons for failure of anticancer drug therapy is development of drug resistance. Anticancer drugs face severe challenges such as poor biodistribution, restricted solubility, inadequate absorption, and drug accumulation. In recent years, “organ-on-a-chip” platforms, which can directly regulate the microenvironment of biomechanics, biochemistry and pathophysiology, have been developed rapidly and have shown great potential in clinical drug research. Lung-on-a-chip (LOC) is a new 3D model of bionic lungs with physiological functions created by micromachining technology on microfluidic chips. This approach may be able to partially replace animal and 2D cell culture models. To overcome drug resistance, LOC realizes personalized prediction of drug response by simulating the lung-related microenvironment in vitro, significantly enhancing therapeutic effectiveness, bioavailability, and pharmacokinetics while minimizing side effects. In this review, we present an overview of recent advances in the preparation of LOC and contrast it with earlier in vitro models. Finally, we describe recent advances in LOC. The combination of this technology with nanomedicine will provide an accurate and reliable treatment for preclinical evaluation.