Centrifugal Chip Research Articles

A fully automated Lab-on-a-Disc platform integrated a high-speed triggered siphon valve for PBMCs extraction

Human Peripheral Blood Mononuclear Cells (PBMCs) are isolated from peripheral blood and identified as any blood cell with a round nucleus that exhibits immune responses and undergoes immunophenotypic changes upon exposure to various pathophysiological stimuli. Obtaining high-recovery and clinical-grade PBMCs without decreasing cell viability and causing stress is crucial for disease diagnosis and successful immunotherapy. However, traditional manual PBMCs extraction methods rely on manual intervention with less recovery rate and reliability. In this study, we introduced a novel and efficient strategy for the fully automated extraction of PBMCs based on a Lab-on-a-Disk (LoaD) platform. The centrifugal chip used percoll as density gradient media (DGM) for separation and extraction on account of the density difference of cells in whole blood, without labeling and any additional extra cellular filtration or cell lysis steps. Above all, we proposed a high-speed triggered siphon valve, which was closed under the speed of cell sedimentation and subsequently opened by increasing speed to complete the extraction of PBMCs. It can avoid the problem that previous siphon valves rely on unstable hydrophilic surface treatment and prime under low/zero speed conditions. With valves and the clock channel integrated on the chip, users can achieve fully automated collection of PBMCs. Compared with the clinical laboratory results, the recovery rate of extracted PBMCs was 80 %. The experimental results prove that the high-speed triggered siphon valve improves the extraction efficiency of PBMCs. The robust chips, which are not only simple to manufacture and assemble but also stable and reliable to use, have great potential in biomedical and clinical applications.

Centrifugal dynamic hybridization conducted in a microfluidic chip for signal enhancement in nucleic acid tests

A rotating platform has been developed using a centrifugal chip holder to mount the standard chips for liquid delivery achieved by centrifugal pumping. This platform allows for dynamic hybridization to be performed in the microchannels constructed in the standard chips made using the 50 mm × 75 mm glass slides which allows for fast hybridization reactions. The results show that when the oligonucleotide-oligonucleotide hybridization is performed, there is good differentiation between perfectly complementary strands over 1 bp-mismatching counterparts when the long target strand is firstly immobilized and the short probe is secondly hybridized (Method 2), but not when the short probe is first immobilized, and the long target is subsequently hybridized (Method 1). When the differentiation between immobilized ginseng PCR product strands is performed, the correct result is achievable by Method 1, after signal enhancement and addition of formamide. The use of Method 2 is successful only when the PCR strand is captured, but not immobilized. In both methods, proper differentiation is achievable using the N1Q probe, un-achievable without centrifugal hybridization.

Fast detection of ß2 microglobulin in patient blood by a handhold centrifugal microfluidic device

Elevating blood ß2 microglobulins (ß2 M) level is one of the indicators of diseases such as diabetes, kidney failure, and tumors. A centrifugal microfluidic chip was designed to conduct latex immunoagglutination assays for testing ß2 M. A handhold centrifuge was assembled to drive solution mixing in the microfluidic chip. An office scanner recorded the binding of ß2 M to the latex beads induced agglutination, reducing reliance on expensive scientific equipment and avoiding environmental light interference. The results revealed that the degree of beads agglutination was proportional to the grayscale value changes (∆G). The ß2 M testing can be accomplished using a 1.25 µL blood sample, and the testing time is 10 min. The limit of detection is 0.0335 mg/L. The recovery rate calculated by the measured and spiked ß2 M concentration ratio is 91.8–111.8%. The centrifugal chip was applied to test 50 patients’ samples. The confusion matrix analyzed the rate of false positives, false negatives, true positives, and true negatives for the ß2 M chip-based test, showing the sensitivity of the ß2 M chip is 77.3%, the specificity is 82.1%, and the efficiency is 80%. The Area Under the Curve analysis shows that the cutoff value (∆G = 1.011) of ∆G in predicting ß2 M elevating has a sensitivity of 100% (95% CI: 92.87% −100%) with the specificity of 92% (95% CI:81.6–96.85%).

Microfluidic compartmentalization to identify gene biomarkers of infection.

Infectious diseases caused by pathogens, such as SARS-COV, H7N9, severe fever with thrombocytopenia syndrome virus, and human immunodeficiency virus, have fatal outcomes with common features of severe fever and subsequent bacterial invasion progressing to multiorgan failure. Gene biomarkers are promising to distinguish specific infections from others with similar presenting symptoms for the prescription of correct therapeutics, preventing pandemics. While routine laboratory methods based on polymerase chain reaction (PCR) to measure gene biomarkers have provided highly sensitive and specific viral detection techniques over the years, they are still hampered by their precision and resource intensity precluding their point-of-care use. Recently, there has been growing interest in employing microfluidic technologies to advance current methods for infectious disease determination via gene biomarker measurements. Here, based on the requirement of infection detection, we will review three microfluidic approaches to compartmentalize gene biomarkers: (1) microwell-based PCR platforms; (2) droplet-based PCR; and (3) point-of-care devices including centrifugal chip, SlipChip, and self-powered integrated microfluidic point-of-care low-cost enabling chip. By capturing target genes in microwells with a small sample volume (∼μl), sensitivity can be enhanced. Additionally, with the advance of significant sample volume minimization (∼pl) using droplet technology, gene quantification is possible. These improvements in cost, automation, usability, and portability have thereby allowed point-of-care applications to decentralize testing platforms from laboratory-based settings to field use against infections.

A portable microfluidic analyzer for integrated bacterial detection using visible loop-mediated amplification

Bacterial contamination has become an important issue for food safety in recent years. Culture-based methods, though a gold standard for bacterial detection, usually take a long time for detection. Microfluidic-based molecular diagnosis can help to achieve rapid and accurate detection of bacteria; however, existing microfluidic systems have limited portability because bulky and complicated instruments are needed. In this study, a portable analyzer was developed to rapidly and simultaneously detect five types of foodborne bacteria (Staphylococcus aureus, Salmonella, Shigella, enterotoxigenic Escherichia coli, and Pseudomonas aeruginosa) on a centrifugal chip. Integration of the processes, including bacteria lysis, visible loop-mediated amplification (LAMP) and detection, was accomplished within 70 min. The analyzer has a compact size (151 × 134 × 110 mm) and two motors positioned on both sides of the centrifugal chip. The upper motor was used to generate a rotating magnetic field for lysing bacteria by bead-beating, whereas the lower motor rotated the chip on demand to execute the following processes: sedimentation of bacteria lysate, mixing of the LAMP buffer and bacteria lysate, and distribution of the mixture to the designated reaction chambers. The LAMP results were detected by a color sensor in the analyzer. Using this portable analyzer, similar detection sensitivities for the five bacteria were achieved compared to that of conventional LAMP in Eppendorf tubes. Considering the portability of the system and its simplicity of operation, the analyzer developed in this study may be of potential use in point-of-care applications.

On-chip ion pair-based dispersive liquid-liquid extraction for quantitative determination of histamine H2 receptor antagonist drugs in human urine

In the present work, an ion-pair based dispersive liquid-liquid microextraction was performed on a centrifugal chip for the first time. The entire DLLME procedure, including flow direction, desperation, and sedimentation of the extracting phase, can be fulfilled automatically on a solitary chip. The chip was made of Poly(methyl methacrylate) (PMMA) and was of two units for two parallel extractions, each consisting of three chambers (for the sample solution, extracting solvents, and sedimentation). As the chip rotated, fluids flowed within the chip, and the dispersion, mixing, extraction, and sedimentation of the final phase were performed on the chip by simply adjusting the spin speed. Determination of two histamine H2 receptor antagonist drugs, cimetidine and ranitidine, as the model analytes from the urine samples was done using the developed on-chip ion-pair based DLLME method followed by an HPLC-UV. The effective parameters on the extraction efficiency of the model analytes were investigated and optimized using the one variable at a time method. Under optimized conditions, the calibration curve was linear in the range of 15–2000 μg L−1 with a coefficient of determination (R2) more than 0.9987. The relative standard deviations (RSD %) for extraction and determination of the analytes were less than 3.7% based on five replicated measurements. LODs less than 10.0 μg L−1 and preconcentration factors higher than 39-fold were obtained for both of the model analytes. The proposed chip enjoys the advantages of both the DLLME method and miniaturization on a centrifugal chip.