Tetraphenylethylene-indole as a novel fluorescent probe for selective and sensitive detection of human serum albumin (HSA) in biological matrices and monitoring of HSA purity and degradation

Human serum albumin (HSA) levels in serum and urine is a crucial biomarker for diagnosing liver and kidney diseases. HSA is used to treat various disorders in clinical practice and as an excipient in the production of vaccine or protein drug, ensuring its purity essential for patient safety. However, selective and sensitive detection of HSA remains challenging due to its structural similarity with bovine serum albumin (BSA) and the inherent complexity of biological matrices. This study presents a novel application of the tetraphenylethylene-indole (TPE-indo) fluorophore for the identification and quantification of HSA. The findings demonstrate that TPE-indo binds specifically to HSA in a 1:1 M ratio, thereby triggering its aggregation-induced emission (AIE) mechanism and producing a selective, sensitive, and rapid "turn-on" fluorescence response. The fluorescence intensity of TPE-indo exhibited minimal interference from proteins, amino acids, sugars, ions, and urine metabolites, and demonstrated a linear correlation with HSA concentration up to 60 μg/mL, with a limit of detection of 0.30 μg/mL. Furthermore, TPE-indo displays a markedly enhanced response to HSA in comparison to BSA, which can be ascribed to the distinct binding modes between TPE-indo and these two proteins. TPE-indo can be used to quantify HSA in serum, grade proteinuria samples, detect BSA adulteration in HSA samples, and real-time monitor HSA degradation processes. This study not only advances the development of efficient HSA detection methods but also highlights the significance of TPE-indo as a versatile tool for bioanalysis and clinical diagnosis.