The Application Of Nanoparticle In Electrogenerated Chemiluminescence Immunoassay

Electrogenerated chemiluminescence (ECL) has received much attention over conventional chemiluminescence because reagents and precursors participating in the reaction are generated in situ when required at an electrode. Electrogenerated chemiluminescence immunoassay ECLIA which uses an ECL species as a label to study the reaction of antigen and antibody and determinate antigen, hapten or antibody has become an increasingly promising immunoassay method owing to its non-radiation, simple instrumentation, high sensitivity, wide dynamic range, reproducibility, simplicity and rapidity analysis, its automation from electrochemical control.This thesis includes a review and a research section.In the review, general introduction to electrogenerated chemiluminescence (ECL) and electrogenerated chemiluminescence immunoassay (ECLIA) including its principle, system and research development were presented. Moreover, the purpose of this research work was presented.The research section contains three subunits. In the research section,We have developed ECL on modified electrode with nanoparticles, moreover ABEI labeled anti-hlgG and luminol-digoxin doped SiC*2 nanoparticles was applied in multilabeled homogeneous ECLIA. The major contents in this thesis are described as follows:1. Study on the electrochemiluminescence behavior of N-(aminobutyl) -N-ethylisoluminol at gold nanoparticles modified graphite electrode Firstly ECL behaviors of at gold nanopartiles modified graphite electrode (GNMGE) in neutral aqueous solution containing hydrogen peroxide were investigated. It was found that, at GNMGE, ECL intensity of ABEI in presence of hydrogen peroxide was dramatically enhanced in neutral aqueous solution, two anodic cyclic voltammetric peaks (cvpl, cvp2) were observed at 0.5 and 0.9V (vs Ag/AgCl), more over the whole current became bigger on gold nanoparticles graphite electrode than those on bare graphite electrode. It demonstrated that gold nanoparticles could catalyze the ABEI in presenceof hydrogen peroxide. The calibration curve for ABEI was obtained from 3.0 X10~-14 to 1.0X 10~-11 mol/L, and the detection limit was 1.0X 10~14 .The relative standard deviation was 1.9% for eleven successive assays at 1.0 X10"11 mol/L ABEI.2. Homogenous electrogenerated chemiluminescence immunoassay using N-(aminobutyl)-N-ethylisoluminol labeled hlgG based on modified gold nanoparticles on graphite electrode It was found that, at modified gold nanoparticles on graphite electrode, ECL intensity of ABEI-labeled anti-hlgG was also dramatically enhanced and ABEI-labeled anti-hlgG also has sensitivity ECL response at this modified electrode. ECL emission was electrochemically generated from the ABEI-labeled anti-hlgG and increased markedly in the presence of antigen due to a more rigid structure of the ABEI moiety. The luminescence intensity of the ABEI-labeled anti-hlgG-hlgG complex exceeded that of ABEI-labeled anti-hlgG itself, so hlgG thus can be sensitivitied determined on GNMGE by subtracting the luminescence intensity of ABEI-labeled anti-hlgG from that of the ABEI-labeled anti-hlgG-hlgG complex. A calibration curve for hlgG and antihlgG was separately from 3.0X10*11 to 1.0X10"9g/niL, Calibration curve of hlgG was obtained from 3.0X10"" to l.OXlO^g/mL. The relative standard deviation was 3.1% and 3.4% for hlgG and anti-hlgG with eleven successive assays at 1.0 X 10'10 g/mL and 1/1000 dilution separately. Sensitivity and accuracy of gold nanoparticles modified graphite electrode for hlgG in human control serum with satisfactory results.3. Homogeneous electrochemiluminescence immunoassay for digoxin antibody and digoxin hapten was developed based on a luminol-digoxin doped SiOj nanopaticles A novel homogeneous electrochemiluminescence immunoassay for digoxin antibody and digoxin hapten was developed using a luminol-digoxin doped SiCh nanopaticles. ECL emission was electrochemically generated from luminol-digoxin doped S1O2 nanopaticles and decreased markedly in the presence of anti-digoxin antibody due to steric hindrance. The anti-digoxin antibody concentration was determined in the range from 1/40000 to 1/167 dilution. Digoxin hapten was determined throughout its therapeutic range with a detection limit of 0.10 ng/mL and in a 3 orders of magnitude range by competitive imrnunoreaction between the digoxin of doped SiO2 nanopaticles and digoxin with fixed quantity of anti-digoxin antibody. The relative standard deviation was 413% and 3.5% for digoxin and anti-digoxin antibody with eleven successive assays at 1.0 X10"9 g/mL and 1/1000 dilution separately. The proposed method has been...