A syringe-based nanoliquid biopsy platform integrating magnetic immunocapture and electrochemical assay for circulating tumor cells enumeration and molecular profiling in NSCLC patients

The number and phenotypic profile of circulating tumor cells (CTCs) have demonstrated their capability in cancer diagnosis and therapeutic efficacy assessment, while programmed death-ligand 1 (PD-L1) and major histocompatibility complex class Ⅱ (MHC-Ⅱ) emerge as core predictive biomarkers for cancer immunotherapy response. Despite their clinical significance, simplified methodologies with potential for point-of-care (POC) adaptation that enable simultaneous detection of CTCs and these two immune-related surface proteins in non-small cell lung cancer (NSCLC) remain scarce, which was quite critical for comprehensive liquid biopsy - based cancer management. In this work, an integrated bioassay for synchronous enumeration of CTCs and quantitative profiling of PD-L1/MHC-Ⅱ expression on captured CTCs from NSCLC patient blood samples was developed. A magnetic immunocapture probe with nanorod architecture was functionalized with dual targeting aptamers, which enabled an enhanced capture efficiency toward CTCs up to 95%. Meanwhile, for differential detection of PD-L1 and MHC-Ⅱ, CdTe and PbS quantum dots (QDs) were conjugated with specific antibodies against PD-L1 and MHC-Ⅱ, respectively, with quantification enabled by electrochemical voltammetric analysis of Cd 2+ and Pb 2+ released from QDs. To simplify assay operation, a disposable syringe filter equipped with nitrocellulose (NC) membrane was employed as the platform for immobilizing antibodies and immuno-recognition. The robustness and efficiency of the integrated system were validated by investigating the correlations between CTC counts, PD-L1/MHC-Ⅱ expression levels, and disease progression in NSCLC patients, which highlighted its potential as a powerful tool for precision immunotherapy guidance.