Rapid point-of-care (POC) quantification of low virus RNA load would significantly reduce the turn-around time for the PCR test and help contain a fast-spreading epidemic. Herein, we report a droplet digital PCR (ddPCR) platform that can achieve this sensitivity and rapidity without bulky lab-bound equipment. The key technology is a flattened pipette tip with an elliptical cross-section, which extends a high aspect-ratio microfluidic chip design to pipette scale, for rapid (<5 min) generation of several thousand monodispersed droplets ∼150 to 350 μm in size with a CV of ∼2.3%. A block copolymer surfactant (polyoxyalkylene F127) is used to stabilize these large droplets in oil during thermal cycling. At this droplet size and number, positive droplets can be counted by eye or imaged by a smartphone with appropriate illumination/filtering to accurately quantify up to 100 target copies. We demonstrate with 2019 nCoV-PCR assay LODs of 3.8 copies per 20 μL of sample and a dynamic range of 4–100 copies. The ddPCR platform is shown to be inhibitor resistant with spiked saliva samples, suggesting RNA extraction may not be necessary. It represents a rapid 1.5-h POC quantitative PCR test that requires just a pipette equipped with elliptical pipette tip, a commercial portable thermal cycler, a smartphone, and a portable trans-illuminator, without bulky and expensive micropumps and optical detectors that prevent POC application.

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Superparamagnetic nanobeads offer several advantages over microbeads for immunocapture of nanocarriers (extracellular vesicles, lipoproteins, and viruses) in a bioassay: high-yield capture, reduction in incubation time, and higher capture capacity. However, nanobeads are difficult to “pull-down” because their superparamagnetic feature requires high nanoscale magnetic field gradients. Here, an electrodeposited track-etched membrane is shown to produce a unique superparamagnetic nano-edge ring with multiple edges around nanopores. With a uniform external magnetic field, the induced monopole and dipole of this nano edge junction combine to produce a 10× higher nanobead trapping force. A dense nanobead suspension can be filtered through the magnetic nanoporous membrane (MNM) at high throughput with a 99% bead capture rate. The yield of specific nanocarriers in heterogeneous media by nanobeads/MNM exceeds 80%. Reproducibility, low loss, and concentration-independent capture rates are also demonstrated. This MNM material hence expands the application of nanobead immunocapture to physiological samples.

Superparamagnetic nanobeads offer several advantages over microbeads for immunocapture of specifc molecular nanocarriers (extracellular vesicles, lipoproteins, and viruses) in a bioassay: high-yield capture, reduction in incuba-tion time and higher capture capacity. However, nanobeads are difficult to “pull down” because their superparamag-netic feature requires high nanoscale magnetic field gradients in addition to high magnetic fields. Here, electroplated track-etched membrane is shown to produce a unique superparamagnetic nanowedge ring with multiple edges around each nanopore. With a uniform external magnetic field, the induced monopole and dipole of this nanowedge junction combine to produce a 10x higher nanobead trapping force. A dense nanobead suspension can be filtered through the magnetic nanoporous membrane (MNM) at a high-throughput with 99% bead capture rate. The capture yield of specific nanocarriers in heterogeneous media (filtered plasma and conditioned cell media) by nanobeads/MNM ex-ceeds 80%. Quantification of RNA cargo in captured extracellular vesicles demonstrate 60x increase in capture rate of a specific microRNA relative to magnetic bead columns. Reproducibility, low loss, and concentration-independent capture rates are also demonstrated. This new MNM material hence significantly expands the application of nano-bead immunocapture to heterogeneous physiological samples, such as blood and saliva, whose molecular analytes are cargoes of nanocarriers.

Superparamagnetic nanobeads offer several advantages over microbeads for immunocapture of specific molecular nanocarriers (extracellular vesicles, lipoproteins, and viruses) in a bioassay: high-yield capture, reduction in incubation time, and higher capture capacity. However, nanobeads are difficult to “pull-down” because their superparamagnetic feature requires high nanoscale magnetic field gradients in addition to high magnetic fields. Here, an electroplated track-etched membrane is shown to produce a unique superparamagnetic nano edge ring with multiple edges around each nanopore. With a uniform external magnetic field, the induced monopole and dipole of this nano edge junction combine to produce a 10x higher nanobead trapping force. A dense nanobead suspension can be filtered through the magnetic nanoporous membrane (MNM) at high throughput with a 99% bead capture rate. The capture yield of specific nanocarriers in heterogeneous media (filtered plasma and conditioned cell media) by nano-beads/MNM exceeds 80%. Quantification of RNA cargo in captured extracellular vesicles demonstrates a 60x increase in the capture rate of a specific microRNA relative to magnetic bead columns. Reproducibility, low loss, and concentration-independent capture rates are also demonstrated. This new MNM material hence significantly expands the application of nanobead immunocapture to heterogeneous physiological samples, such as plasma and saliva.