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UBC Theses and Dissertations

Cellulose-based biosensors of human neutrophil elastase (HNE) toward chronic wound point-of-care diagnostics Saisuwan, Ravi


Chronic wounds, which fail to heal or heal only very slowly, remain a major challenge in medical treatment and a burden to healthcare systems. Chronic wounds are exacerbated by bacterial infection and endogenous proteases. In particular, increased levels of human neutrophil elastase (HNE) have been observed in chronic wound fluid, and has been utilized as a severity indicator via clinical assays. To facilitate chronic wound treatment, a facile, in situ, detection method for HNE would be advantageous. Here, cellulose-based analytical devices have been designed and produced as a proof-of-concept for chronic wound point-of-care diagnostics. Specifically, two distinct fluorogenic HNE substrates amenable to click-chemistry were synthesized based on tetrapeptide (Ala4) conjugates to the rhodamine derivatives, carboxyrhodamine110-PEG3-azide (cRho110-PEG₃-N₃) and rhodamine 110 (Rho110). Michaelis-Menten kinetics were used to demonstrate activity of HNE toward these compounds that was comparable to known chromogenic substrates. Following attachment under mild aqueous conditions to alkyne-functionalized Whatman No. 1 filter paper, a pure cellulosic substrate and model for cotton gauze, HNE detection on a solid surface was demonstrated visually under specific illumination and was quantified with a fluorescence scanner. These results validate the concept of in situ protease detection using modified cellulose surfaces to monitor chronic wounds toward improved treatment outcomes. Furthermore, the modular design of the cellulose-based analytical devices presented here suggests a broader potential for the detection of specific protease activity in diverse applications.

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