Multiplexed detection of SARS-CoV-2 based on upconversion luminescence nanoprobe/MXene biosensing platform for COVID-19 point-of-care diagnostics.

Multiplexed detection is essential in biomedical sciences since it is more efficient and accurate than single-analyte detection. For an accurate early diagnosis of COVID-19, a multiplexed detection strategy is required to avoid false negatives with the existing gold standard assay. Nb2CTx nanosheets were found to efficiently quench the fluorescence emission of lanthanide-doped upconversion luminescence nanoparticles at wavelengths ranging from visible to near-infrared spectrum. Using this broad-spectrum quencher, we developed a label-free FRET-based biosensor for rapid and accurate detection of SARS-CoV-2 RNA. To target ORF and N genes, two types of oligo-modified lanthanide-doped upconversion nanoparticles can be used simultaneously to identify-two sites in one assay via upconversion fluorescence enhancement intensity measurement with detection limits of 15 pM and 914 pM, respectively. Moreover, with multisite cross-validation, this multiplexed and sensitive biosensor is capable of simultaneous and multicolor analysis of two gene fragments of SARS-CoV-2 Omicron variant within minutes in a single homogeneous solution, which significantly improves the detection efficiency. The diagnosis result via our assay is consistent with the PCR result, demonstrating its application in the rapid and accurate screening of multiple genes of SARS-CoV-2 and other infectious diseases.

Rapid point-of-care detection of SARS-CoV-2 RNA with smartphone-based upconversion luminescence diagnostics.

Accurate COVID-19 screening via molecular technologies is still hampered by bulky instrumentation, complicated procedure, high cost, lengthy testing time, and the need for specialized personnel. Herein, we develop point-of-care upconversion luminescence diagnostics (PULD), and a streamlined smartphone-based portable platform facilitated by a ready-to-use assay for rapid SARS-CoV-2 nucleocapsid (N) gene testing. With the complementary oligo-modified upconversion nanoprobes and gold nanoprobes specifically hybridized with the target N gene, the luminescence resonance energy transfer effect leads to a quenching of fluorescence intensity that can be detected by the easy-to-use diagnostic system. A remarkable detection limit of 11.46 fM is achieved in this diagnostic platform without the need of target amplification, demonstrating high sensitivity and signal-to-noise ratio of the assay. The capability of the developed PULD is further assessed by probing 9 RT-qPCR-validated SARS-CoV-2 variant clinical samples (B.1.1.529/Omicron) within 20 min, producing reliable diagnostic results consistent with those obtained from a standard fluorescence spectrometer. Importantly, PULD is capable of identifying the positive COVID-19 samples with superior sensitivity and specificity, making it a promising front-line tool for rapid, high-throughput screening and infection control of COVID-19 or other infectious diseases.