Study On The Detection Of Thiols And DNA Based On Chemiluminescence And Photoelectrochemical Techniques

Glutathione (GSH) is a major low molecular weight thiols in body fluids, which is abundant in many kinds of tissues and cells. It is important in the many biological processes such as human metabolism, intracellular redox equilibrium and protein synthesis. Deoxyribonucleic acid (DNA) is known as inherited substance in body fluids. Much more effort have been paid on the detection of specific DNA sequences and DNA mutation because it is significant for early diagnosis and cure for disease.This paper focused on the detection of DNA and GSH using chemiluminescence and photoelectrochemical techniques. We developed a new style chemiluminescence probe and photoelectrochemical DNA biosensor for the detection of GSH and DNA. It provided a newly developed method for clinic diagnoses of many diseases at early stage. The major contents of this paper are as follows:1. We provide a dicopper complex CL probe for the determination of GSH and related cellular thiols for the first time. The structure of Cu₂L₂ was characterized by Elemental, 1H NMR and MS analysis. The UV and visible absorption spectra showed that GSH was able to form complexes with copper (II) derived from probe Cu₂L₂. This competitive complexation with copper (II) may lead to the destruction of the peroxidase-like dicopper complex Cu₂L₂, then decreases in CL intensity were occurred accordingly. The relatively low detection limit of 1.8 nM and a broad dynamic range of 8.0×10^-9 2.0×10^-7 M were achieved. The developed CL assay based on CL probe Cu₂L₂ was considerably attractive for rapid, sensitive and selective detection of cellular thiols. The novel strategy provides an alternative method for molecule recognition using CL probe.2. A novel photoelectrochemical biosensor for detection of GSH was developed based on thiol-disulfide exchange reaction. photoelectrochemically active species [Ru(bpy)₂dppz](BF4)₂·2H₂O was synthesized and [Ru(bpy)₂dppz]²⁺-doped SiO₂ nanoparticles were prepared by water-in-oil (W/O) mode. The ITO electrodes which have many hydroxyl groups on the surface could be silanized in an ethanol solvent containing 3-MPTES. The surface was then immersed in PDEA solution, and disulfide bridge was formed. Finally, ITO electrode was combined with [Ru(bpy)₂dppz]²⁺-doped SiO₂ nanoparticles through the interaction of glutaraldehyde. In our experiment, the detection limit of GSH was 7.8×10^-8 mol/L (3σ) under the optimum conditions. In addition, we investigate the electrode of stability and recovery, the result was satisfactory.3. Here, CL and photoelectrochemstry were combined in one system firstly. Thus a simple electrochemical analyzer can work obviating the use of physical light source involved in conventional photoelectrochemistry assays. The attachment involves the formation of a monolayer of 3-aminopropyltriethoxysilane (APS) on SnO₂ nanoparticle modified ITO electrode. Then the surface is modified further with a cross-linker, glutaraldehyde, by means of a schiff base reaction. The left aldehyde group can be used to covalent immobilization of amine-capped capture DNA. After hybridization with target DNA and intercalation with Ru(bpy)₂dppz²⁺, the target DNA can be quantitatively detected when the photoelectrochemically active species rutheniumbipyridine derivatives was excited by CL reaction of CIPO-H₂O₂-9,10-diphenylanthrancene outside. By using this novel method, target DNA could be detected conveniently as as well as other bio-active molecules. The selected CL system can also used to excied AuNPs-Ru(bpy)₂dppz²⁺ complex modified on ITO electrode as well as (PDDA/CdS)5 multilayers modified on planar gold electrode and produce preferable photocurrent.