Confocal Microscopy Analysis of Condensate Fluorescence at Varying Tyrosinase Concentrations.

Varying concentrations of tyrosinase (0.005, 0.01, 0.025, 0.05, 0.075, 0.1, 0.15, 0.2, and 0.5 mg mL⁻¹) were added to peptide/RNA condensates (2 mM/1 mg mL⁻¹) at pH 7.5 and 37 °C. Each sample was imaged using a laser scanning confocal microscope (as described above) at either λ_ex = 405 nm or λ_ex = 640 nm following 10 min incubation with the enzyme. Each data point represents the average of 10 condensates, and SDs are presented. The data were fitted to a four-parameter logistic regression with iteration algorithm of Orthogonal Distance Regression and Levenberg Marquardt for λ_ex = 405 nm and λ_ex = 640 nm, respectively (Eq. 3). A1 and A2 are the initial and final fluorescence intensity, respectively, x is the tyrosinase concentration, x_1/2 is the x value that results in a 50% fluorescence intensity response, and p is the power. The LOD was calculated as the initial value (A1) plus three times the SD (Table 3).

Sigmoidal fitting, linear regression