Monitoring the level of matrix metalloproteinase-9 (MMP-9) and inhibiting its expression is important for the diagnosis and treatment of various diseases. However, the analysis of MMP-9 is challenging owing to its very low content in the blood, especially at the early stages of diseases. Therefore, we developed an ultrasensitive and easy-to-use immunosensor based on a three-dimensional (3D) bioplatform for the determination of the total MMP-9 concentration in plasma. The used 3D bioplatform (G2 poly(amidoamine) dendrimer; PAMAM) improved the sensitivity of the determination by significantly expanding the surface area of the receptor layer. The antigen-antibody recognition process was controlled by quartz crystal microbalance with dissipation (QCM-D) and electrochemical impedance spectroscopy (EIS). The effect of the orientation of antibody molecules in the sensing layer on the work parameters of the immunosensor was analyzed using unmodified PAMAM (PAMAM-NH2) and PAMAM functionalized with -COOH groups (PAMAM-COOH). The developed immunosensor based on PAMAM-NH2 was characterized by a lower detection limit (LOD = 2.0 pg⋅mL-1) and wider analytical range (1·10-4 - 5 μg⋅mL-1 for EIS and QCM-D) compared to PAMAM-COOH immunosensor (EIS: 1·10-4 - 0.5 μg⋅mL-1; QCM-D: 5·10-4 - 0.5 μg⋅mL-1). The functionality of the proposed device was verified in spiked plasma. The recoveries determined in commercial human and rat plasma and noncommercial rat plasma were very close to the value of 100% and in the range of 96-120% for Au/PAMAM-NH2/Ab and Au/PAMAM-COOH/Ab immunosensors, respectively. The designed analytical devices showed high selectivity and sensitivity without the use of any amplifiers such as metal nanoparticles or enzymes.
Keywords: Cancer biomarker; Electrochemical immunosensor; Gravimetric detection; Matrix metalloproteinase 9; PAMAM bioplatform.
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