Konkanavalin A Baskılanmış Yüzey Plazmon Rezonans Temelli Biyosensörün Hazırlanması

Abstract

Within the scope of this thesis, a Concanavalin A imprinted surface plasmon resonance (SPR) biosensor was prepared for the determination of Concanavalin A. At the beginning, the surface of the SPR biosensor chip was modified using allyl mercaptan. Then, a pre-polymerization complex was generated utilizing N-methacryloyl-(L)-histidine methyl ester (MAH) monomer in various molar ratios with the Concanavalin A template molecule. The pre-complex, HEMA as the functional monomer, was added to the polymer mixture prepared using EGDMA as the crosslinker and polymerized using UV light on the surface of the SPR chip in the presence of the AIBN initiator. The same methods were repeated without utilizing the Concanavalin A template molecule as a control, and a non-imprinted SPR biosensor based on Concanavalin A was created. Atomic force microscopy (AFM), ellipsometry, and contact angle measurements were used to characterize the manufactured SPR chips. pH scanning was used to establish the optimal pH value for the Concanavalin A molecule using Concanavalin A solutions made with pH 4, 5, 6, and 7.4 phosphate buffers and a Concanavalin A suppressed biosensor. The sensorgrams obtained indicated that the proper pH for the Concanavalin A molecule iv is 7.4 phosphate buffer. The sensitivity of the SPR biosensor was then tested using kinetic analysis with Concanavalin A solutions ranging from 0.01 to 50 ng/mL. The accuracy of the imprinted biosensor created from sensorgrams derived from the measurement data was 99 percent in the concentration range (0.01-5 ng/mL); it was measured with 96 percent accuracy in the concentration range (10-50 ng/mL). The limit of detection (LOD) was determined to be 0.0038 ng/mL at the limit of quantification (LOQ) of 0.0011 ng/mL. The binding kinetics of Concanavalin A were determined using solutions produced at various concentrations. The Langmuir was proven to be the best appropriate adsorption model for the developed biosensor. Selectivity researches were conducted using SPR biosensors that were either suppressed or not suppressed with Concanavalin A, as well as competing molecules such as bovine serum albumin (BSA) and myoglobin (Myg), which have a similar molecular structure. Concanavalin A was isolated from soybean for use in real sample studies. The kinetic analysis of samples made from the genuine sample was carried out using the Concanavalin A suppressed SPR biosensor, and the quantity of Concanavalin A and recovery were determined using the sensorgrams obtained. To evaluate the Concanavalin A molecule's suitability for use in the artificial control plasma environment, kinetic analyses of Concanavalin A solutions prepared at various concentrations in the artificial control plasma were performed using the suppressed SPR biosensor, and the amount of Concanavalin A and recovery was calculated using the sensorgrams obtained. Four sequential analyses were performed on the same day to determine the reusability of the Concanavalin A molecule. Additionally, kinetic analyses were conducted at 1, 2, 4, and 6 months to determine the biosensor's applicability at various time points. Concanavalin in its prepared state A suppressed SPR biosensor has been shown to be a sensitive, rapid, long-lasting, and cost-effective method for determining Concanavalin A.