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On-site severe acute respiratory syndrome coronavirus 2 (SARS CoV-2) serological assays allow for timely in-field decisions to be made regarding patient status, also enabling population-wide screening to assist in controlling the coronavirus disease 2019 (COVID-19) pandemic. Here we propose a rapid microfluidic serological assay with two unique functions of nanointerstice filling and digitized flow control, which enable the fast/robust filling of the sample fluid as well as precise regulation of duration and volume of immune reaction. Developed microfluidic assay showed enhanced limit of detection, and 91.67% sensitivity and 100% specificity (n = 152) for clinical samples of SARS CoV-2 patients. The assay enables daily monitoring of IgM/IgG titers and patterns, which could be crucial parameters for convalescence from COVID-19 and provide important insight into how the immune system responds to SARS CoV-2. The developed on-site microfluidic assay presented the mean time for IgM and IgG seroconversions, indicating that these titers plateaued days after seroconversion. The mean duration from day 0 to PCR negativity was 19.4 days (median 20 d, IQR 16–21 d), with higher IgM/IgG titres being observed when PCR positive turns into negative. Simple monitoring of these titres promotes rapid on-site detection and comprehensive understanding of the immune response of COVID-19 patients. • Rapid on-site SARS CoV-2 serological assays allow for timely in-field decisions. • Microfluidic serological assay with nanointerstices and digitized flow control. • Enhances the immune reaction and the limit of detection. • Platform shows 91.67% sensitivity and 100% specificity for SARS CoV-2. • Enables daily monitoring of IgM/IgG titers and patterns. [ABSTRACT FROM AUTHOR]

Co-circulation of coronavirus disease 2019 (COVID-19) and dengue fever has been reported. Accurate and timely multiplex diagnosis is required to prevent future pandemics. Here, we developed an innovative microfluidic chip that enables a snapshot multiplex immunoassay for timely on-site response and offers unprecedented multiplexing capability with an operating procedure similar to that of lateral flow assays. An open microchannel assembly of individually engineered microbeads was developed to construct nine high-density test lines, which can be imaged in a 1 mm2 field-of-view. Thus, simultaneous detection of multiple antibodies would be achievable in a single high-resolution snapshot. Next, we developed a novel pixel intensity-based imaging process to distinguish effective and non-specific fluorescence signals, thereby improving the reliability of this fluorescence-based immunoassay. Finally, the chip specifically identified and classified random combinations of arbovirus (Zika, dengue, and chikungunya viruses) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antibodies within 30 min. Therefore, we believe that this snapshot multiplex immunoassay chip is a powerful diagnostic tool to control current and future pandemics. • A bead-based one-step multiplex immunoassay platform is developed. • Operating procedure is similar to that of lateral flow assay kits. • Open microfluidic-based high-density self-assembly of bead test lines is presented. • An imaging process facilitates simultaneous and reliable detection in a single snapshot. • Multiplex immunoassay is demonstrated with various mixture of four antibodies including anti-SARS-CoV-2 RBD. [ABSTRACT FROM AUTHOR]

Abstract In the study, we describe an oscillatory flow-assisted efficient target enrichment method by using a particle-based microarray device. Periodic oscillating flow effectively increased the mixing and binding performance between the target molecules in the sample solution and surface functionalized microparticles. Particles were trapped, secured, and released with an elastic microvalve structure operated via differences in the flow conditions. Single particle (20-µm diameter) trapping efficiency exceeded 95%. Secured particles can freely move inside each array element based on oscillating sample flow. Furthermore, the particles can be released from the array and collected at the outlet of the device, and this provides an opportunity for further off-chip analysis. As a proof-of-concept, we used the interaction between streptavidin-coated microparticles and fluorescence labeled biotin solution and demonstrated that target enrichment and detection based on oscillatory flow were significantly more efficient than that based on unidirectional or static flow. The applicability of the method was further examined by conducting an on-chip immunoassay to detect the presence of anti-Zika nonstructural protein 1 (NS1) monoclonal antibody. The limit of detection (LOD) was as low as 1 ng/mL with an assay time of only 10 min and less than 10 µL of sample consumption. Highlights • Particle-based microarray device is demonstrated through a structure deformation. • Oscillatory flow-assisted efficient target enrichment method is presented. • Assay is performed with small sample volume (< 10 µL) and in a short time (10 min). • Limit of detection (LOD) for detecting anti-Zika NS1 mAb is as low as 1 ng/mL. [ABSTRACT FROM AUTHOR]

• Simple microfluidic sandwich immunoassay was designed to be operated by sample injection only for point of care testing • Closing fluid vent by PC-connected small equipment made the flow pause to provide sufficient reaction time for immunoassay • Limit of detection was significantly increased by pausing the flow • Developed assay showed same limit of detection with benchtop sandwich immunoassay as well as 10 times higher than commercial rapid kit Microfluidic immunoassays have gained immense importance as a point-of-care testing (POCT) platform owing to their low reagent volume requirements, low cost, portability, and rapid reaction. However, they are characterized by inconsistency and lack of reproducibility. In this study, we developed a microfluidic immunoassay platform with a simple signal enhancement technique using fluid vent control. The components of this fluorescence-based immunoassay were successfully pre-loaded in the microfluidic device. The analytes reacted with the detection antibody conjugated on fluorescence beads, and bound to capture the antibody immobilized zone during channel flow. The fluid vent was closed using a small PC-connected equipment when the fluid reached the target displacement. The pausing of the fluid provided sufficient time for the immune reaction, and this addition of the fluid pausing significantly enhanced the immune reaction and increased the limit of detection (LOD) of the immunoassay. The LOD of the developed assay was the same as that of the sandwich fluorescence immunoassay in terms of quantity. Furthermore, the LOD was significantly higher than that of the commercial rapid kits. Therefore, our simple technique could be employed for cost-effective and accurate immunoassays in the field of POCT. [ABSTRACT FROM AUTHOR]