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

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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. Graphical Abstract

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