General characteristics of isothermal amplification assay for SARS-CoV-2 detection.

Method	Required enzyme	Probes number	Temperature (°C)	Time (min)	Target	LOD (copies per μL)	Sensitivity	Advantagesa	Disadvantagesa	Ref.
NEAR	Nicking endonuclease, strand-displacing DNA	2	60–65	5–15	RdRp gene	0.125	48–71.7%	Fast testing, universally in various environments	Low sensitivity	149
NASBA	RNase H, reverse transcriptase, T7 DNA-dependent RNA polymerase	2	41	90–120	S gene N gene	0.5	98.15–100%	High sensitivity	Required RNase inhibitors	150
Non-enzymatic isothermal strand displacement and amplification	Restriction endonucleases, strand-substituting DNA polymerases	3	42	<30	RdRp gene, N gene	10	96.77%	No RNA reverse transcription step, fast testing	Non-specific reactions/false positives	151
Rolling circle amplification	DNA ligase, DNA polymerase	8	30–37	<120	N gene S gene	1	—	High specificity	False negatives and false positives	5

^aAdvantages and disadvantages are compared to RT-qPCR methods.