RNA-extraction-free nano-amplified colorimetric test for point-of-care clinical diagnosis of COVID-19.
Nat Protoc
; 16(6): 3141-3162, 2021 06.
Article
in English
| MEDLINE | ID: covidwho-1209962
ABSTRACT
The global pandemic of coronavirus disease 2019 (COVID-19) highlights the shortcomings of the current testing paradigm for viral disease diagnostics. Here, we report a stepwise protocol for an RNA-extraction-free nano-amplified colorimetric test for rapid and naked-eye molecular diagnosis of COVID-19. The test employs a unique dual-prong approach that integrates nucleic acid (NA) amplification and plasmonic sensing for point-of-care detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), with a sample-to-assay response time of <1 h. The RNA-extraction-free nano-amplified colorimetric test utilizes plasmonic gold nanoparticles capped with antisense oligonucleotides (ASOs) as a colorimetric reporter to detect the amplified nucleic acid from the COVID-19 causative virus, SARS-CoV-2. The ASOs are specific for the SARS-CoV-2 N-gene, and binding of the ASOs to their target sequence results in the aggregation of the plasmonic gold nanoparticles. This highly specific agglomeration step leads to a change in the plasmonic response of the nanoparticles. Furthermore, when tested using clinical samples, the accuracy, sensitivity and specificity of the test were found to be >98.4%, >96.6% and 100%, respectively, with a detection limit of 10 copies/µL. The test can easily be adapted to diagnose other viral infections with a simple modification of the ASOs and primer sequences. It also provides a low-cost, rapid approach requiring minimal instrumentation that can be used as a screening tool for the diagnosis of COVID-19 at point-of-care settings in resource-poor situations. The colorimetric readout of the test can even be monitored using a handheld optical reader to obtain a quantitative response. Therefore, we anticipate that this protocol will be widely useful for the development of biosensors for the molecular diagnostics of COVID-19 and other infectious diseases.
Full text:
Available
Collection:
International databases
Database:
MEDLINE
Main subject:
RNA, Viral
/
Oligonucleotides, Antisense
/
Metal Nanoparticles
/
COVID-19 Nucleic Acid Testing
/
SARS-CoV-2
/
COVID-19
/
Gold
Type of study:
Diagnostic study
/
Prognostic study
Limits:
Humans
Language:
English
Journal:
Nat Protoc
Year:
2021
Document Type:
Article
Affiliation country:
S41596-021-00546-w
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