CD274 and hepatocellular carcinoma: For the evaluation of mRNA expression levels in HCC CTCs, qRT‒PCR analysis was conducted on samples containing 0, 50, 100, and 1000 CTCs, with relative quantification performed via the 2−ΔΔCt method.[7, 48] Figure 2d summarizes the relative expression levels of EpCAM, GPC3, and PD‐L1 mRNAs across different CTC counts.