Fig. 5: Simultaneous detection of multiple clinically relevant biomarkers across multiple molecular classes in water and 20% human serum matrices.

Reactions were incubated at 37 °C for 3 h. a Schematic of membrane-less protocell array setup for multimodal, simultaneous detection of diverse classes of clinically relevant biomarkers. Purple and yellow circles indicate micro-basins containing zinc and vitamin B₁₂ sensors, respectively. Red and blue circles indicate micro-basins containing Stx1 (S1) and Stx2 (S2) toehold switches, respectively. b Representative fluorescence image (top) and quantification of the representative image and its replicates (bottom), demonstrating multimodal target measurement using protocell arrays with simultaneous detection of ion, small molecule, RNA, and DNA targets from the same bulk phase. Data for each biological replicate are provided in Supplementary Fig. 11. c Multimodal target detection in a 20% human serum matrix. Representative fluorescence image (top) and quantification of the representative image and its replicates (bottom), demonstrating the robustness of protocell arrays to a complex sample matrix. The asterisk (*) next to the zinc concentration indicates the total zinc concentration is 2 µM in the bulk phase after accounting for the zinc remaining in chelated human serum (Supplementary Fig. 14). Of note, the sensor plasmid concentrations were increased in c compared to b to yield comparable output signal; this tuning was necessary due to the decrease in protein expression that occurs in human serum⁵. Data for each biological replicate are provided in Supplementary Fig. 13. Data are presented as mean values ± SD of 9 replicates (3 biological replicates × 3 technical replicates). Hollow circles indicate the mean of each biological replicate. Scale bar is 1 mm. Details on CFE lysates used, plasmid concentrations, and reaction additives are provided in Supplementary Table 1.