scTP-seq: 用于NSCLC的单细胞转录组与细胞内蛋白同步分析技术

Abstract

scTP-seq enables simultaneous profiling of transcriptomes and intracellular proteins in unfixed single cells. • It combines droplet microfluidics with antibody-oligonucleotide conjugates for high-throughput multi-omics. • scTP-seq reveals protein-defined states and tumor-suppressive genes missed by transcriptome-only approaches. • This platform provides a streamlined, scalable solution for protein-informed discovery in complex tissues. Single-cell RNA sequencing has revolutionized cell typing but cannot concurrently profile intracellular proteins, creating a critical blind spot that obscures post-transcriptional regulation and impedes functional biomarkers discovery. To address this, we developed scTP-seq (single-cell T ranscriptome and P rotein seq uencing), a platform that enables simultaneous single-cell profiling of the transcriptome and intracellular proteins without prior cell manipulation, ensuring minimal technical bias. We validated scTP-seq against 10× Genomics data for PBMCs and a published NSCLC transcriptome atlas, confirming high transcriptional concordance (R 2 values ranging from 0.814 to 0.989). Applied to NSCLC, scTP-seq uncovered a protein-defined CYFRA21-1-high alveolar epithelial type II cell subpopulation that was transcriptionally indistinguishable, demonstrating its power to reveal hidden cellular states. By correlating T cell transcripts with protein levels of CD276, FGL-1, and IL-4I1, we identified low-abundance tumor-suppressive genes that are overlooked by scRNA-seq. Leveraging this protein-guided resolution, we dissected the immunosuppressive niche, pinpointing exhausted CD8+ T cells as a clinical hub interacting with cDC2 cells via dense checkpoints including SPP1-CD44 and NECTIN2-TIGIT. Collectively, scTP-seq resolves the critical blind spot in single-cell analysis by enabling a protein-informed discovery pipeline, positioning it as a powerful tool for uncovering latent therapeutic targets and advancing cancer immunology.