Rapid One-Step Detection of Viral Particles Using an Aptamer-Based Thermophoretic Assay

Abstract

Rapid and sensitive identification of viral pathogens such as SARS-CoV-2 is a critical step to control the pandemic disease. Viral antigen detection can compete with gold-standard PCR-based nucleic acid diagnostics in terms of better reflection of viral infectivity and reduced risk of contamination from enzymatic amplification. Here, we report the development of a one-step thermophoretic assay using an aptamer and polyethylene glycol (PEG) for direct quantitative detection of viral particles. The assay relies on aptamer binding to the spike protein of SARS-CoV-2 and simultaneous accumulation of aptamer-bound viral particles in laser-induced gradients of temperature and PEG concentration. Using a pseudotyped lentivirus model, a limit of detection of ∼170 particles μL −1 (26 fM of the spike protein) is achieved in 15 min without the need of any pretreatment. As a proof of concept, the one-step thermophoretic assay is used to detect synthetic samples by spiking viral particles into oropharyngeal swabs with an accuracy of 100%. The simplicity, speed, and cost-effectiveness of this thermophoretic assay may expand the diagnostic tools for viral pathogens.

Functional nucleic acids as modular components against SARS-CoV-2: From diagnosis to therapeutics

“…The assay platform has potential value for POC with rapid response and good portability. In another fascinating study, Deng et al (2021) instead combined a thermoelectrophoretic approach for rapidly quantification of pseudo-SARS-CoV-2 viral particles due to the high-affinity aptamer of SARS-CoV-2 S protein. With the merits of rapidity (15 min), high sensitivity (detection limit of 176 TU μL −1 ), high accuracy (100%), one-step detection (no pre-treatment) and low cost (US$ 0.4 per test), this method has the capacity to be a validated tool for large-scale population-based screening towards SARS-CoV-2 infection.…”

Overcoming the limitations of COVID-19 diagnostics with nanostructures, nucleic acid engineering, and additive manufacturing

“…In addition to PGMs and LFA, DNA aptamers have been integrated to a wide variety of technologies to detect different pathogens and proteins, ranging from portable fluorescence [101] , [102] , [103] and colorimetric assays [104] , [105] , to rapid assays that require instrumentation such as MST [106] . Given the progress made so far, we are confident that in the near future DNA aptamers specific to SARS-CoV-2 will be interfaced with new technologies summarized in this review to make it possible for portable, on-site, and real-time detection of infectious SARS-CoV-2.…”

High‐Affinity Dimeric Aptamers Enable the Rapid Electrochemical Detection of Wild‐Type and B.1.1.7 SARS‐CoV‐2 in Unprocessed Saliva

“…[ 33 , 34 , 35 , 36 , 37 , 38 ] However, all aptamers reported to date show affinity similar to antibodies ( K d ranges from 2–85 nM), and as such only a few aptamer‐based rapid saliva tests for SARS‐CoV‐2 have been reported to date. [ 28 , 39 ] We hypothesized that these issues could be solved from two approaches: (1) deriving a spike‐binding DNA aptamer with extremely high affinity as the molecular recognition element, and (2) utilizing an extremely sensitive signal transduction strategy to report the viral binding by the aptamer. This rational approach was expected to overcome the loss of sensitivity reported for patient saliva samples, resulting in a rapid, simple and sensitive test for detecting SARS‐CoV‐2 from human saliva without amplification.…”