Study Of Immunosensors Based On Various Nanocomposites And Nanopaticles For The Detection Of Prostate Specific Antigen

BackgroundBiosenors are widely used for the analysis of biological analytes. Biosenors have become a very active research topic, owing to the advantages of their high sensitivity, fast analytical time, simple measurement, low-cost and high selectivity. At present, much attention has been devoted to the development of the third-generation electrochemical biosensor based on direct electrochemistry. The nanomaterials provide a novel way to construct the third-generation biosensor. Nanoparticles with larger specific interfaceareas, desirable biocomapatibility, and a high surface free energy could adsorb more antibodies without the loss of their biological activities, which amplifed sensitivity of the amperometric immunosensors. It has been reported that the crucial aspect in the fabrication of a biosensor is the immobilization of various nanocomposites and nanopaticles in high amounts with retention of their bioactivity onto the electrode surface.ObjectiveTo establish a sensitive and strict-specificity amperometric immunosensors which can detect the prostate specific antigen (PSA) based on various nanocomposites and nanopaticles. And to optimize possible experimental conditions affecting the efficiency of electron-transfer processes, then to characterize electrochemical characteristics of the modified electrode with cyclic voltammetry (CV). At last clinical samples are detected and the results are compared with those detected by conventional immunoassay in order to explore the feasibility of the detection of clinical samples.Materials and methods1. Prepare indium tin oxides (ITO) by successive sonication in acetone, ethanol doubly and distilled water .2. Multi-wall carbon nanotubes (MWNT) ultrasonically cleaned in concentrated nitric acid, then rinsed with water and baked dry.3. Prepare chitosan-prussian blue nanocomposite (CS-PB), Au nanoparticles (Aunano) , Au nanoparticles -prussian blue nanocomposite (Aunano- PB) and N- (2-aminoethyl) -3- aminopropyltrimethoxysilance - Dextran-Fe3O4 composite magnetic nanoparticles (AEAPS- Dextran-Fe3O4).4. Investigate the size and morphology of nanoparticles conducted with Transmission electron micrographs (TEMs) and Ultra violet/visible (UV–vis) spectroscopy. Determination of Fe3O4 and dextran from AEAPS/Dextran-Fe3O4 by atomic absorption and anthrone-sulfuric acid colorimetry method.5. Prepare immunosensors of BSA/anti-PSA/ Aunano /CS-PB/MWNT/ DpAu/ITO and BSA/anti- PSA / Aunano / AEAPS-Dextran-Fe3O4/ Aunano-PB /ITO.6. Characterize electrochemical characteristics of the modified electrode with cyclic voltammetry (CV) in a 0.1 M phosphate buffer solution (pH 7.0). To improve the sensitivity, detection limit of amperometric immunosensors the experimental conditions were optimized.7. Characterize electrochemical characteristics of the modified electrode with interference test, stability test, repeatability test, recovery test and contrast test.Results1. Using ultrasonic cleaning technique, we have purified ITO.2. After ultrasonically cleaning in concentrated nitric acid, we have purified MWNT which were functionalized with carboxylic acid groups.3. Found a method to prepare CS-PB nanocomposite, Aunano, Aunano -PB nanocomposite and AEAPS-Dextran-Fe3O4 composite magnetic nanoparticles. The observation of transmission electron microscope (TEM) showed that the diameter of these monodisperse nanoparticles ranged from 16 to 25 nm. Investigations of optical properties of nanoparticles were also conducted with Ultra violet/visible (UV–vis) spectroscopy.4. By such optimization strategies as experimental condition, we have constructed a kind of amperometric immunosensor which could detect the PSA, then established the corresponding technical protocol. And the amperometric immunosensors could complete the detection in 10 min with the higher sensitivity, wider concentration ranges.5. The results are compared with that conventional immunoassay, and the correlation coefficients are good, which showed that the amperometric immunosensors could be used for clinical detection.ConclusionsTwo amperometric immunosensors established in this study for the determination of PSA have developed by means of self-assembled monolayers (SAMs) and deposition method onto a ITO electrode based on CS-PB nanocomposite, MWNT, Aunanoor Aunano-PB nanocomposite, AEAPS-Dextran-Fe3O4 composite magnetic nanoparticles, Aunano as matrixes. Therefore, the platform that combines the advantages of nanostructured materials open the doors to the new and exciting possibilities for the development of immunosensor using different transduction modes.