Duplex Electrochemical Microfluidic Sensor for COVID-19 Antibody Detection: Natural versus Vaccine-Induced Humoral Response.

The rapid transmission and resilience of coronavirus disease 2019 (COVID-19) have led to urgent demands in monitoring humoral response for effective vaccine development, thus a multiplex co-detection platform to discriminate infection-induced from vaccine-induced antibodies is needed. Here a duplex electrochemical immunosensor for co-detection of anti-nucleocapsid IgG (N-IgG) and anti-spike IgG (S-IgG) is developed by using a two-working electrode system, via an indirect immunoassay, with antibody quantification obtained by differential pulse voltammetry. The screen-printed electrodes (SPEs) are modified by carbon black and electrodeposited gold nanoflowers for maximized surface areas, enabling the construction of an immunological chain for S-IgG and N-IgG electrochemical detection with enhanced performance. Using an optimized immunoassay protocol, a wide linear range between 30-750 and 20-1000 ng mL-1 , and a limit of detection of 28 and 15 ng mL-1 are achieved to detect N-IgG and S-IgG simultaneously in serum samples. This duplex immunosensor is then integrated in a microfluidic device to obtain significantly reduced detection time (≤ 7 min) while maintaining its analytical performance. The duplex microfluidic immunosensor can be easily expanded into multiplex format to achieve high throughput screening for the sero-surveillance of COVID-19  and other infectious diseases.

Glioblastoma adhesion in a quick-fit hybrid microdevice.

Translational research requires reliable biomedical microdevices (BMMD) to mimic physiological conditions and answer biological questions. In this work, we introduce a reversibly sealed quick-fit hybrid BMMD that is operator-friendly and bubble-free, requires low reagent and cell consumption, enables robust and high throughput performance for biomedical experiments. Specifically, we fabricate a quick-fit poly(methyl methacrylate) and poly(dimethyl siloxane) (PMMA/PDMS) prototype to illustrate its utilities by probing the adhesion of glioblastoma cells (T98G and U251MG) to primary endothelial cells. In static condition, we confirm that angiopoietin-Tie2 signaling increases the adhesion of glioblastoma cells to endothelial cells. Next, to mimic the physiological hemodynamic flow and investigate the effect of physiological electric field, the endothelial cells are pre-conditioned with concurrent shear flow (with fixed 1 Pa shear stress) and direct current electric field (dcEF) in the quick-fit PMMA/PDMS BMMD. With shear flow alone, endothelial cells exhibit classical parallel alignment; while under a concurrent dcEF, the cells align perpendicularly to the electric current when the dcEF is greater than 154 V m- 1. Moreover, with fixed shear stress of 1 Pa, T98G glioblastoma cells demonstrate increased adhesion to endothelial cells conditioned in dcEF of 154 V m- 1, while U251MG glioblastoma cells show no difference. The quick-fit hybrid BMMD provides a simple and flexible platform to create multiplex systems, making it possible to investigate complicated biological conditions for translational research.

Flow of wormlike micellar solutions around microfluidic cylinders with high aspect ratio and low blockage ratio.

We employ time-resolved flow velocimetry and birefringence imaging methods to study the flow of a well-characterized shear-banding wormlike micellar solution around a novel glass-fabricated microfluidic circular cylinder. In contrast with typical microfluidic cylinders, our geometry is characterized by a high aspect ratio α = H/W = 5 and a low blockage ratio ß = 2r/W = 0.1, where H and W are the channel height and width, and the cylinder radius r = 20 µm. The small cylinder radius allows access up to very high Weissenberg numbers 1.9 ≤ Wi = λMU/r ≤ 3750 (where λM is the Maxwell relaxation time) while inertial effects remain entirely negligible (Reynolds number, Re < 10-4). At low Wi values, the flow remains steady and symmetric and a birefringent region (indicating micellar alignment and tensile stress) develops downstream of the cylinder. Above a critical value Wic ≈ 60 the flow transitions to a steady asymmetric state, characterized as a supercritical pitchfork bifurcation, in which the fluid takes a preferential path around one side of the cylinder. At a second critical value Wic2 ≈ 130, the flow becomes time-dependent, with a characteristic frequency f0 ≈ 1/λM. This initial transition to time dependence has characteristics of a subcritical Hopf bifurcation. Power spectra of the measured fluctuations become complex as Wi is increased further, showing a gradual slowing down of the dynamics and emergence of harmonics. A final transition at very high Wic3 corresponds to the re-emergence of a single peak in the power spectrum but at much higher frequency. We discuss this in terms of possible flow-induced breakage of micelles into shorter species with a faster relaxation time.

Microcontact printing with aminosilanes: creating biomolecule micro- and nanoarrays for multiplexed microfluidic bioassays.

Microfluidic systems integrated with protein and DNA micro- and nanoarrays have been the most sought-after technologies to satisfy the growing demand for high-throughput disease diagnostics. As the sensitivity of these systems relies on the bio-functionalities of the patterned recognition biomolecules, the primary concern has been to develop simple technologies that enable biomolecule immobilization within microfluidic devices whilst preserving bio-functionalities. To address this concern, we introduce a two-step patterning approach to create micro- and nanoarrays of biomolecules within microfluidic devices. First, we introduce a simple aqueous based microcontact printing (µCP) method to pattern arrays of (3-aminopropyl)triethoxysilane (APTES) on glass substrates, with feature sizes ranging from a few hundred microns down to 200 nm (for the first time). Next, these substrates are integrated with microfluidic channels to then covalently couple DNA aptamers and antibodies with the micro- and nanopatterned APTES. As these biomolecules are covalently tethered to the device substrates, the resulting bonds enable them to withstand the high shear stresses originating from the flow in these devices. We further demonstrated the flexibility of this technique, by immobilizing multiple proteins onto these APTES-patterned substrates using liquid-dispensing robots to create multiple microarrays. Next, to validate the functionalities of these microfluidic biomolecule microarrays, we perform (i) aptamer-based sandwich immunoassays to detect human interleukin 6 (IL6); and (ii) antibody-based sandwich immunoassays to detect human c-reactive protein (hCRP) with the limit of detection at 5 nM, a level below the range required for clinical screening. Lastly, the shelf-life potential of these ready-to-use microfluidic microarray devices is validated by effectively functionalizing the patterns with biomolecules up to 3 months post-printing. In summary, with a single printing step, this aminosilane patterning technique enables the creation of functional microfluidic micro- and nano-biomolecule arrays, laying the foundation for high-throughput multiplexed bioassays.

In situ pressure measurement within deformable rectangular polydimethylsiloxane microfluidic devices.

In this paper, we present a simple procedure to incorporate commercially available external pressure transducers into existing microfluidic devices, to monitor pressure-drop in real-time, with minimal design modifications to pre-existing channel designs. We focus on the detailed fabrication steps and assembly to make the process straightforward and robust. The work presented here will benefit those interested in adding pressure drop measurements in polydimethylsiloxane (PDMS) based microchannels without having to modify existing channel designs or requiring additional fabrication steps. By using three different devices with varying aspect ratio channels ([Formula: see text], width/depth), we demonstrate that our approach can easily be adapted into existing channel designs inexpensively. Furthermore, our approach can achieve steady state measurements within a matter of minutes (depending on the fluid) and can easily be used to investigate dynamic pressure drops. In order to validate the accuracy of the measured pressure drops within the three different aspect ratio devices, we compared measured pressure drops of de-ionized water and a 50 wt. % glycerol aqueous solution to four different theoretical expressions. Due to the deformability of PDMS, measured pressure drops were smaller than those predicted by the rigid channel theories (plate and rectangular). Modification of the rigid channel theories with a deformability parameter α provided better fits to the measured data. The elastic rectangular expression developed in this paper does not have a geometric restriction and is better suited for microchannels with a wider range of aspect ratios.