A hyaluronidase detection platform combined with enzyme-responsive hydrogel and periodic magnetic-modulated electrochemical analysis system

Abstract

Frequent invasive cystoscopies are required for bladder cancer management due to high recurrence and progression rates, significantly affecting the quality of life for patients. Hyaluronidase is a promising urinary biomarker that could potentially reduce the reliance on cystoscopy; however, its clinical utility depends on highly sensitive and specific detection methods. A major challenge is the common presence of hematuria in bladder cancer patients, as red blood cells in urine substantially interfere with hyaluronidase assays. Here, we present a platform for quantifying urinary hyaluronidase activity that operates directly in urine without the need for sample pre-treatment. This platform employs a hyaluronic acid hydrogel capable of specifically recognizing hyaluronidase. Enzymatic degradation of the hydrogel releases embedded magnetic beads, effectively converting the enzyme activity into a quantifiable magnetic bead signal. These released magnetic beads are then determined using a developed periodic magnetic-modulated electrochemical analysis system. The platform achieves hyaluronidase activity detection in the range of 2–25 U/mL, with a limit of detection of 0.414 U/mL. The specificity of hyaluronic acid hydrogel degradation, combined with the magnetic separation of magnetic beads, effectively reduces interference from other urinary components. In addition, the periodic magnetic controlled strategy introduces frequency-domain analysis into electrochemical amperometric signal processing, thereby avoiding the interference of baseline drift. Importantly, the platform demonstrates robust performance in detecting hyaluronidase in artificial hematuria samples, underscoring its substantial potential for clinical application in non-invasive bladder cancer monitoring.