A lab-on-a-chip for the concurrent electrochemical detection of SARS-CoV-2 RNA and anti-SARS-CoV-2 antibodies in saliva and plasma.

Rapid, accurate and frequent detection of the RNA of SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) and of serological host antibodies to the virus would facilitate the determination of the immune status of individuals who have Coronavirus disease 2019 (COVID-19), were previously infected by the virus, or were vaccinated against the disease. Here we describe the development and application of a 3D-printed lab-on-a-chip that concurrently detects, via multiplexed electrochemical outputs and within 2 h, SARS-CoV-2 RNA in saliva as well as anti-SARS-CoV-2 immunoglobulins in saliva spiked with blood plasma. The device automatedly extracts, concentrates and amplifies SARS-CoV-2 RNA from unprocessed saliva, and integrates the Cas12a-based enzymatic detection of SARS-CoV-2 RNA via isothermal nucleic acid amplification with a sandwich-based enzyme-linked immunosorbent assay on electrodes functionalized with the Spike S1, nucleocapsid and receptor-binding-domain antigens of SARS-CoV-2. Inexpensive microfluidic electrochemical sensors for performing multiplexed diagnostics at the point of care may facilitate the widespread monitoring of COVID-19 infection and immunity.

Laboratory-Generated DNA Can Cause Anomalous Pathogen Diagnostic Test Results.

The coronavirus disease 2019 (COVID-19) pandemic has brought about the unprecedented expansion of highly sensitive molecular diagnostics as a primary infection control strategy. At the same time, many laboratories have shifted focus to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) research and diagnostic development, leading to large-scale production of SARS-CoV-2 nucleic acids that can interfere with these tests. We have identified multiple instances, in independent laboratories, in which nucleic acids generated in research settings are suspected to have caused researchers to test positive for SARS-CoV-2 in surveillance testing. In some cases, the affected individuals did not work directly with these nucleic acids but were exposed via a contaminated surface or object. Though researchers have long been vigilant of DNA contaminants, the transfer of these contaminants to SARS-CoV-2 testing samples can result in anomalous test results. The impact of these incidents stretches into the public sphere, placing additional burdens on public health resources, placing affected researchers and their contacts in isolation and quarantine, removing them from the testing pool for 3 months, and carrying the potential to trigger shutdowns of classrooms and workplaces. We report our observations as a call for increased stewardship over nucleic acids with the potential to impact both the use and development of diagnostics. IMPORTANCE To meet the challenges imposed by the COVID-19 pandemic, research laboratories shifted their focus and clinical diagnostic laboratories developed and utilized new assays. Nucleic acid-based testing became widespread and, for the first time, was used as a prophylactic measure. We report 15 cases of researchers at two institutes testing positive for SARS-CoV-2 on routine surveillance tests, in the absence of any symptoms or transmission. These researchers were likely contaminated with nonhazardous nucleic acids generated in the laboratory in the course of developing new SARS-CoV-2 diagnostics. These contaminating nucleic acids were persistent and widespread throughout the laboratory. We report these findings as a cautionary tale to those working with nucleic acids used in diagnostic testing and as a call for careful stewardship of diagnostically relevant molecules. Our conclusions are especially relevant as at-home COVID-19 testing gains traction in the marketplace and these amplicons may impact on the general public.

Utilising Commercially Fabricated Printed Circuit Boards as an Electrochemical Biosensing Platform.

Printed circuit boards (PCBs) offer a promising platform for the development of electronics-assisted biomedical diagnostic sensors and microsystems. The long-standing industrial basis offers distinctive advantages for cost-effective, reproducible, and easily integrated sample-in-answer-out diagnostic microsystems. Nonetheless, the commercial techniques used in the fabrication of PCBs produce various contaminants potentially degrading severely their stability and repeatability in electrochemical sensing applications. Herein, we analyse for the first time such critical technological considerations, allowing the exploitation of commercial PCB platforms as reliable electrochemical sensing platforms. The presented electrochemical and physical characterisation data reveal clear evidence of both organic and inorganic sensing electrode surface contaminants, which can be removed using various pre-cleaning techniques. We demonstrate that, following such pre-treatment rules, PCB-based electrodes can be reliably fabricated for sensitive electrochemical biosensors. Herein, we demonstrate the applicability of the methodology both for labelled protein (procalcitonin) and label-free nucleic acid (E. coli-specific DNA) biomarker quantification, with observed limits of detection (LoD) of 2 pM and 110 pM, respectively. The proposed optimisation of surface pre-treatment is critical in the development of robust and sensitive PCB-based electrochemical sensors for both clinical and environmental diagnostics and monitoring applications.

Integrated Electrochemical Biosensors for Detection of Waterborne Pathogens in Low-Resource Settings.

More than 783 million people worldwide are currently without access to clean and safe water. Approximately 1 in 5 cases of mortality due to waterborne diseases involve children, and over 1.5 million cases of waterborne disease occur every year. In the developing world, this makes waterborne diseases the second highest cause of mortality. Such cases of waterborne disease are thought to be caused by poor sanitation, water infrastructure, public knowledge, and lack of suitable water monitoring systems. Conventional laboratory-based techniques are inadequate for effective on-site water quality monitoring purposes. This is due to their need for excessive equipment, operational complexity, lack of affordability, and long sample collection to data analysis times. In this review, we discuss the conventional techniques used in modern-day water quality testing. We discuss the future challenges of water quality testing in the developing world and how conventional techniques fall short of these challenges. Finally, we discuss the development of electrochemical biosensors and current research on the integration of these devices with microfluidic components to develop truly integrated, portable, simple to use and cost-effective devices for use by local environmental agencies, NGOs, and local communities in low-resource settings.

A PNA-based Lab-on-PCB diagnostic platform for rapid and high sensitivity DNA quantification.

We report the development of a Lab-on-PCB DNA diagnostic platform, exploiting peptide nucleic acid (PNA) sequences as probes. The study demonstrates the optimization and characterization of two commercial PCB manufacturing gold electroplating processes for biosensing applications. Using an optimized ratio of PNA with a spacer molecule (MCH), the lowest limit of detection (LoD) to date for PCB-based DNA biosensors of 57 fM is reported. The study also showcases a fully integrated Lab-on-PCB microsystem designed for rapid detection, which employs PCB-integrated sample delivery, achieving DNA quantification in the 0.1-100 pM range for 5 µL samples analyzed within 5 min under continuous flow. The demonstrated biosensor proves the capability of PCB-based DNA biosensors for high sensitivity and paves the way for their integration in Lab-on-PCB DNA diagnostic microsystems.