High Sensitivity Extended Nano-Coulter Counter for Detection of Viral Particles and Extracellular Vesicles

Abstract

We present a chip-based extended nano-Coulter counter (XnCC) that can detect nanoparticles affinity-selected from biological samples with low concentration limit-of-detection that surpasses existing resistive pulse sensors by 2–3 orders of magnitude. The XnCC was engineered to contain 5 in-plane pores each with an effective diameter of 350 nm placed in parallel and can provide high detection efficiency for single particles translocating both hydrodynamically and electrokinetically through these pores. The XnCC was fabricated in cyclic olefin polymer (COP) via nanoinjection molding to allow for high-scale production. The concentration limit-of-detection of the XnCC was 5.5 × 103 particles/mL, which was a 1,100-fold improvement compared to a single in-plane pore device. The application examples of the XnCC included counting affinity selected SARS-CoV-2 viral particles from saliva samples using an aptamer and pillared microchip; the selection/XnCC assay could distinguish the COVID-19­(+) saliva samples from those that were COVID-19(−). In the second example, ovarian cancer extracellular vesicles (EVs) were affinity selected using a pillared chip modified with a MUC16 monoclonal antibody. The affinity selection chip coupled with the XnCC was successful in discriminating between patients with high grade serous ovarian cancer and healthy donors using blood plasma as the input sample.

Unraveling RNA Conformation Dynamics in Mitochondrial Encephalomyopathy, Lactic Acidosis, and Stroke-like Episode Syndrome with Solid-State Nanopores

“…The early work that established the foundations for the nanopore methods include sensing polymers in solution; use of protein nanopores to sequence DNA; − monitor enzyme kinetics; detect damaged DNA; discriminate between polymers based on their size; − discriminate between different metal nanoparticles; detect, quantify, and discriminate between different proteins; − and discriminate between a native protein/point mutants in the same/and post-translational modifications . Additionally, important examples of foundational work on solid-state pores also includes detection of genomic DNA, viral particles, and polysaccharides. − Among these applications, nanopores have shown high promises in identifying the intricate dynamics of RNA in general, − and tRNA in particular, offering a pathway to explore their structural variations that have functional significance.…”

Characterization of Extracellular Vesicles by Resistive-Pulse Sensing on In-Plane Multipore Nanofluidic Devices

“…Resistive-pulse sensing − is a convenient method for characterizing the size, shape, and charge of single particles and has been used to characterize EV size and shape. ,,, One approach to fabricate resistive-pulse devices is to pattern the micro- and nanofluidic channels in the plane of the substrate. , These in-plane nanochannels and nanopores are typically a set of interconnected nanoscale trenches fabricated directly by focused ion beam (FIB) milling into glass substrates − or replication of nanoscale features into polymers, which, in turn, are sealed with a cover plate. With the in-plane architecture, multiple pores in series, , along with other fluidic elements, e.g., filters and reactors, are easily coupled in a single device without the need to align components after fabrication.…”