Figure 2
From: Field-deployable, rapid diagnostic testing of saliva for SARS-CoV-2
(A) HP-LAMP assay workflow. Heat-inactivated saliva (5 µl) was added to each of the two 1.5 ml microcentrifuge tubes pre-filled with the reaction mixture, incubated at 63°C for 30 min and then visualized for colorimetric change (yellow = positive; red = negative). At least one out of two tubes must turn yellow to interpret the assay result as positive. The minimum equipment needed to run the assay is a disposable transfer pipette, 2 heat blocks, and microcentrifuge tubes prefilled with reaction mixture. No prior RNA extraction or treatment is required. (B) Genome map showing targeted region of primers used for HP-LAMP in green. Locations of primers and probes from the CDC 2019-nCoV Real-Time RT-PCR Diagnostic Panel/New York SARS-CoV-2 Real-time RT-PCR Diagnostic Panel, and the New England Biolabs’ (NEB) SARS-CoV-2 assay are indicated in Red. Whereas previously used primers and probes targeted the GC-rich regions located at the 5′ and 3′ ends of the viral RNA, the primers used for HP-LAMP target the central portion of the viral RNA, which is better protected from digestion by salivary exonucleases. The targeted region of the HP-LAMP primer set with alignments of other Betacoronavirus genomes are featured29,15. Each nucleotide is shown (A: green; G: gray; T: red; C: blue). The percentage of GC-content across the genome is indicated (http://genome.ucsc.edu)30,16. (C) Determining the limit of detection (LoD) of the HP-LAMP assay. The concentrations indicated show copies of heat-inactivated SARS-CoV-2 per µl of saliva. NC = negative control with no SARS-CoV-2 added. ‘ + ’ = Positive HP-LAMP result; ‘-’ = Negative HP-LAMP result. The color of each box is taken directly from its corresponding reaction tube. (D) Cross-reactivity of HP-LAMP assay on common pathogens. Inactivated known respiratory pathogens (n = 21) along with inactivated SARS-CoV-2 virus were tested using HP-LAMP assay. All pathogens showed negative detection results in HP-LAMP assay, expect for SARS-CoV-2 virus. (E) Representative results of HP-LAMP testing on clinical samples. (F) Table shows positive and negative percentage agreement of HP-LAMP for detection of SARS-CoV-2 in saliva compared with nasopharyngeal swab RT-PCR results.
