Nanoparticles Amplified QCM Sensor for Enzyme Activity Evaluation
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Nanoparticles Amplified QCM Sensor for Enzyme Activity Evaluation M. Stoytcheva, R. Zlatev, G. Montero, B. Valdez, M. Schorr Presenting author’s email: [email protected] Engineering Institute of the Autonomous University of Baja California, Mexicali, Mexico ABSTRACT This investigation introduces a new very simple and efficient approach for QCM sensor response amplification, developed for hydrolases activity determination. For this purpose, the QCM crystal surface was modified with nanoparticles loaded enzyme substrate. During the enzymatic substrate degradation, the heavier nanoparticles were also released from the sensitive layer together with the substrate degradation products. Nanoparticles removal resulted in QCM signal amplification due to the higher nanoparticles specific mass compared with the specific mass of the substrate. The suggested concept was successfully applied for creating of simple biosensing platforms for trypsin and lipase activity determination in real time using respectively SiO2 nanoparticles loaded olive oil and Ag nanoparticles loaded gelatin as enzyme substrates. Up to 10 times amplification of the QCM signal was reached applying the proposed approach compared with the common one. INTRODUCTION Trypsin (EC 3.4.21.4) is a protein degrading enzyme of the group of the serine proteases employed in some technological processes of the food and beverage industries to improve the workability of dough, in tenderizing of meat, in production of protein hydrolysates, and during cold stabilization of beer, among other [1]. Trypsin is used in biochemistry for development of cell and tissue culture protocols [2-4], and for protein identification through peptide sequencing techniques [5]. In medicine, trypsin activity serves as a specific and reliable diagnostic parameter of pancreatic function and its alteration: cystic fibrosis, chronic pancreatitis, etc. [6, 7]. The industrial, biotechnological, and biochemical importance of trypsin, as well as its clinical significance generated a great interest in developing of methods for quantification of its activity. Spectrophotometric and sensitive radioimmunoassay-based techniques [8, 9] are mainly employed, as well as temperature, pressure [10], and humidity [11] change measurement due to the action of trypsin on gelatin films. Numerous (bio)sensors-based methods were developed recently as well. Chuang et al. [12] exploited the surface plasmon resonance wavelength shift of gelatin modified colloidal gold nanoparticles when they aggregate after gelatin digestion to quantify trypsin activity. Zaccheo et al. [13] reported a self-powered galvanic cell-based sensor for nakedeye detection of serum trypsin. Series of more sensitive and faster electrochemical sensors for trypsin activity determination were developed by Biloivan et al. [14] Ionescu et al. [15, 16], Fordyce et al. [17], Adjemian et al. [18], Baş et al. [19], Stoytcheva et al. [20], Chen et al. [6] with conductometric, amperometric, voltammetric, and potentiometric detection. Krause et al. [21] and St
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