A dual-mode RNA-splitting aptamer biosensor for sensitive HIV Tat peptide detection via colorimetry and fluorescence.

Early diagnosis of human immunodeficiency virus (HIV) is critical for effective treatment; however, traditional antibody methods encounter challenges during the infection window, and nucleic acid tests require specialized equipment. In this study, a dual-mode ribonucleic acid (RNA)-splitting aptamer biosensor was developed to target the HIV trans-activator of transcription (Tat) protein, a key HIV biomarker for viral replication throughout the infection cycle. The biosensor integrates colorimetric and fluorescent detection techniques by utilizing gold nanoparticles (AuNPs) and two types of aptamers, one labeled with carboxyfluorescein (FAM). In the presence of Tat, RNA-splitting aptamers adsorb onto AuNPs, protecting them from recombination, while the fluorescence of FAM is quenched via fluorescence resonance energy transfer (FRET). Aptamers form a ternary complex with Tat, preventing adsorption and leading to thioflavin T-induced aggregation of AuNPs, accompanied by a visible color change and fluorescence signal restoration. The biosensor demonstrated excellent sensing performance, with a linear range of 0.5-60 nM and a detection limit of 0.28 nM, successfully detecting Tat in human serum. Therefore, this low-cost dual-mode detection platform offers a promising tool for early HIV diagnosis and potential applications in clinical and point-of-care fields.