Fig. 3

Optimization of experimental parameters of CHA process. A The principle of the CHA process. B Varying concentrations of initial H1 and H2 (5 nM, 10 nM, 15 nM, 20 nM, and 25 nM) with fixed ratio at 1:1. C Varying ratios between H1 and H2: 1:0.5, 1:1, 1:1.5, and 1:2, with fixed H1 concentration at 15 nM. D Varying incubation temperatures from 25 to 44 °C with optimized concentration and ratio. E Varying incubation time length for CHA amplification (30–120 min) with optimized conditions. All error bars represent mean ± s.d., n = 3