Fusion of Molecular and Mechanical Phenotypes Enables High-Purity and Low-Loss Reacquisition of Viable CTCs for Transcriptome Analysis

Abstract

The high-purity and low-loss reacquisition of viable circulating tumor cells (CTCs) is crucial for enabling downstream omics analysis of CTCs and currently represents key challenges limiting their application in clinical diagnosis and pathological research. Given the limitations of traditional methods that rely solely on a molecular or mechanical phenotype for CTCs acquisition, this study introduces an innovative approach that fuses the inherent molecular and mechanical phenotypes of CTCs into a new mechanical phenotype, thereby achieving high-purity preconcentration and low-loss reacquisition of CTCs. Specifically, CTCs in blood are immunomodified using calcium carbonate microspheres (CCMSs) conjugated with antibodies, transforming the molecular phenotype (membrane protein expression) into an additional mechanical phenotype (increased size and reduced deformability). This transformation enhances the mechanical phenotype distinctions between CTCs and white blood cells, enabling high-purity preconcentration of CTCs on a single-cell trapping array chip. Since CCMSs can be reversibly eliminated under weak acid, captured CTCs can be nondestructively reacquired with 93.10% in microliter-scale solution, allowing for subsequent omics analysis. In a breast cancer mouse model, the counts and transcriptome analysis of CTCs provide valuable insights into assessing tumor occurrence and progression.