Fig. 4
Linearity studies of HCC EV Digital Scoring Assay. A Linearity studies of HCC EV Digital Scoring Assay using synthetic plasma sample, prepared by serially spiking HepG2 EVs into the EV-depleted HD plasma. EV Click Beads were utilized to enrich HepG2 EVs in conjunction with the cocktail of the 3 TCO-antibodies targeting EpCAM, CD147, and ASGPR1. RT-dPCR was performed to obtain the absolute quantification of 12 HCC EV-specific gene candidates. B Robust linearity correlations were identified between the concentration of spiked HepG2 EVs and measured mRNA copy numbers for all 12 HCC EV-specific gene candidates over the concentration range of 0 – 6 × 109 HepG2 EVs per µL, with R-squared values ranging from 0.991 to 0.999. ASGPR1, asialoglycoprotein receptor 1; EpCAM, epithelial cellular adhesion molecule; EV, extracellular vesicle; HCC, hepatocellular carcinoma; HD, healthy donor; RT-dPCR, reverse-transcription digital PCR; TCO, trans-cyclooctene