Fig 1.
Performance of single primer sets.
Each primer set was assayed with 24 repeats with 50 copies of SARS-CoV-2 RNA template per reaction. Each primer set was assigned a sensitivity score: sensitive (*** 16–24 positives); medium (** 9–15 positives) or poor (* 1–8 positives). (A) RT-LAMP reactions conducted with M1800 Colorimetric LAMP at 65°C. (B) With M1800 at 60°C. (C) With E1700 LAMP Mix at 65°C. (D) With E1700 at 60°C.
Fig 2.
Increased detection sensitivity by combining 2 primer sets.
Reactions were performed with single primer sets and various combinations of 2 primer sets in the presence of 25 copies of SARS-CoV-2 RNA template per reaction. The sensitivity score was adjusted as: sensitive (*** 12–24 positives); medium (** 6–11 positives) or poor (* 1–5 positives). (A) Reactions with single primer sets were compared to combinations of 2 sets using M1800 at 65°C. (B) With M1800 at 60°C. (C) Comparing reactions with 4 different dual primer set combinations (S10+S11, S10+S12, S11+S12 and N2+E1) in M1800 at 65 and 60°C. (D) The same dual primer sets in E1700 at 65 and 60°C.
Fig 3.
Effect of combining 2 primer sets in the presence of extremely low SARS-CoV-2 RNA template.
A similar test was designed as Fig 2 but using only ~12.5 copies of SARS-CoV-2 RNA template per reaction. (A) Single primer set reactions for primers S10 and S11 versus a combination of them in M1800 at 65 and 60°C. (B) In E1700 at 65 and 60°C. (C) Single primer set reactions for primers N2 and E1 versus a combination of them in M1800 at 65 and 60°C. (D) In E1700 at 65 and 60°C. Sensitivity Score for 12.5 copies: *** 12–24 positives; ** 6–11 positives; * 1–5 positives.
Fig 4.
Effect of combining 3 primer sets.
Detection sensitivity by a combination of 2 primer sets was compared with that by a combination of 3 primer sets in the presence of ~12.5 copies of template RNA per reaction in M1800 and E1700 at 65 and 60°C. (A) Combinations of 2 primer sets for S10, S11 and S12 versus a combination of all 3 sets using M1800 at 65 and 60°C. (B) S10, S11 and S12 in E1700 at 65 and 60°C. (C) Combinations of 2 primer sets for N2, E1 and As1e versus a combination of all 3 sets using M1800 at 65 and 60°C. (D) N2, E1 and As1e in E1700 at 65 and 60°C. Sensitivity Score for 12.5 copies: *** 12–24 positives; ** 6–11 positives; * 1–5 positives.