Fig. 4

Measurement of GPC1 mRNA in pancreatic cancer cell-derived EVs and patient serum EVs. a TIRF images of GPC1 mRNA expression in AsPC-1 EVs (upper row, signal) and HPDE6-C7 EVs (bottom row, control) using LN–MB, LN–CHDC, LPHN–MB, and LPHN–CHDC, respectively. b Fluorescence intensity of AsPC-1 EVs (signal) and HPDE6-C7 EVs (control) with the same concentration around 10⁸ mL⁻¹ treated with LN–MB, LN–CHDC, LPHN–MB, and LPHN–CHDC, respectively. c Fluorescence signal amplification capability of LN–CHDC, LPHN–MB, or LPHN–CHDC relative to LN–MB based on AsPC-1 EV-associated fluorescence. d Signal-to-background ratios of LN–MB, LN–CHDC, LPHN–MB, and LPHN–CHDC (signal represents fluorescence intensity of AsPC-1 EVs; background represents fluorescence intensity of HPDE6-C7 EVs). e TIRF images of AsPC-1 EVs with 10-, 50-, 250-, and 1000-fold dilution detected by LPHN–CHDC. EV concentration for each dilution measured by NanoSight LM10 was ~10⁷ mL⁻¹, ~2 × 10⁶ mL⁻¹, ~4 × 10⁵ mL⁻¹, and ~10⁵ mL⁻¹, respectively. f Calibration curve for fluorescence intensity of GPC1 mRNA expression in AsPC-1 EVs using LPHN–CHDC vs. low concentration of EV. The LOD of AsPC-1 EVs with LPHN–CHDC was 57,550 per mL. g Representative TIRF images of GPC1 mRNA expression in serum EVs of discovery cohort, healthy donors (n = 60), BPD patients (n = 15), stage I–II pancreatic cancer patients (n = 86), and stage III–IV pancreatic cancer patients (n = 32), total n = 193, using LPHN–CHDC (upper). Fluorescence intensities of GPC1 mRNA expression calculated by METLAB in the discovery cohort (bottom) (paired two-tailed Student’s t test, ****P < 0.0001). h C _t value for GPC1 mRNA in serum EVs of discovery cohort (paired two-tailed Student’s t test, ****P < 0.0001). i ROC curve analysis of discovery cohort. j Representative TIRF images of GPC1 mRNA expression in serum EVs of validation cohort, healthy donors (n = 15), BPD patients (n = 8), stage I–II pancreatic cancer patients (n = 25), and stage III–IV pancreatic cancer patients (n = 23), total n = 71, using LPHN–CHDC (upper). Fluorescence intensities of GPC1 mRNA expression calculated by METLAB in the validation cohort (bottom) (paired two-tailed Student’s t test, ****P < 0.0001). k ROC curve analysis of validation cohort. Data represent mean ± s.d., n = 3, three technical replicates