Ultrasensitive Multiplexed Detection of Protein Biomarkers via Aptamer-Driven T7 RNA Polymerase Amplification

作者：LI Yong-jun, Zixin Yang, Jiahao Wu, Liye Chen, Qian Zhao, Haowen Huang · 发表于：Analytical Chemistry · 年份：2025 · DOI：10.1021/acs.analchem.5c04805 · 被引用次数：4 · 研究领域：Advanced biosensing and bioanalysis techniques、Biosensors and Analytical Detection、Molecular Biology Techniques and Applications

Accurate and multiplexed detection of protein biomarkers in complex biological fluids is essential for early diagnosis and precision medicine. However, existing technologies face critical trade-offs among sensitivity, specificity, multiplexing capability, and operational simplicity. Antibody-based assays like ELISA often lack the sensitivity for early detection, while nucleic acid amplification methods are poorly suited for protein targets and require complex workflows. This creates a pressing need for a simple, highly sensitive, and interference-free approach for simultaneous protein detection in real-world samples. Here, we report an aptamer-driven T7 RNA polymerase cascade amplification system for visual, dual-channel detection of protein biomarkers directly in serum. The platform converts target recognition into isothermal RNA transcription, producing red or green fluorescence via fluorogenic RNA-dye complexes. Using alpha-fetoprotein and human serum albumin as models, the system achieved femtogram-per-milliliter sensitivity, excellent specificity, and broad linear ranges. A red-green fluorescence logic enables intuitive clinical interpretation. Validation with spiked and clinical samples showed recovery rates between 90 and 110% and strong agreement with ELISA results. These findings demonstrate that protein-responsive nucleic acid circuits can achieve precise, multiplexed detection with instrument-free signal amplification. By integrating molecular recognition, signal a...