Figure 4. Application of fluorescent biosensor to detect 2′, 3′-cGAMP in mammalian cell extracts.
(A) Schematic representation of the experimental procedure for cell extract analysis using the fluorescent biosensor. (B, C) Fluorescence readings for different concentrations of L929 (B) or HEK293T (C) cell extracts (DNA- or mock-stimulated). (D) Fluorescence readings for L929 cell extracts (DNA- or mock-stimulated) treated with active or denatured SVPD prior to biosensor addition. (E) Standard curve of fluorescence readings from mock-stimulated L929 cell extracts doped with known concentrations of 2′, 3′-cGAMP (black circles). Average fluorescence reading from DNA-stimulated L929 cell extracts is shown in open circle. Error bars indicate standard deviations for three (B, C) or two (D, E) independent biological replicates. P-values are calculated from student’s t-test comparison: **, P < 0.01; ***, P < 0.001, n.s, not significant, P > 0.05.