Based On Nano Composite Material For The Detection Of Active Substances In Vivo By Electrochemical Immunosensors

Biosensor is a kind of special sensor, which is to identify the bioactive substances(hormones, enzymes, cells, nucleic acids, etc.) and convert the signal into a physical signal, namely light, electricity and other signal which were detected and analyzed by special instruments. It has the characteristics of high specificity, high sensitivity and low cost. The biosensor can monitor continuously in the complex system. Owning to easily realizing automation, miniaturization, therefore, it can be widely used in the field of clinical diagnosis(such as p H, blood gas analyzer, etc.),biological sample analysis, food safety, military, environmental monitoring and so on.As for biosensor, it is an important way to choose suitable surface modification materials for improvement of the analysis of biological sensors. Nano materials have been widely used in biosensors, because of its unique physical and chemical properties(good ductility, good biocompatibility and excellent conductivity), In particular, it has a large specific surface area and being modified easily. So it can increase the amount of biological molecules. thereby it can significantly improve the sensitivity of the biosensor. Based on the advantages of biosensors andnano materials, this work constructs two kinds of electrochemical sensor with nano composite modified for the detection of bioactive substances in vivo(take prolactin for example). detailed work is as follows:1 Based on graphene/chitosan/Au nanoparticles modified electrodes detection of prolactin hormoneProlactin is one of the important hormones in human body. Prolactin levels may be of some significance in distinguishing between epilepsy and non epileptic. In this study, a novel amperometric immunosensor for detection of prolactin has been developed using the glassy carbon electrodes modified with graphene and electrochemically deposited gold nanoparticles.The hybrid nanocomposites not only offer a appropriate microenvironment for immobilization of primary antibodies and maintain their biological activity, but also accelerate electron transfer between the immobilized proteins and electrode substrates. In the presence of prolactin, a sandwich-type immune complex was formed on the sensing interface.Finally, the streptavidin-alkaline phosphatase was bound to the biotinylated secondary antibodies, catalyzing the hydrolysis of the substrate α-naphthyl phosphate. Under optimal conditions, the immunosensor exhibited a linear response to prolactin from 100 pg m L-1 to 50 ng m L-1,with a detection limit of 38.9 pg m L-1(S/N=3). And it has good specificity and reproducibility and it was successful applied in the detection of human serum prolactin.2 Electrochemical detection of prolactin by using graphene/single walled carbon nanotubes/chitosan/Au nanoparticles and antibody coated gold nanoparticlesA glassy carbon electrode(GCE) was modified with a hybrid material consisting of graphene, single walled carbon nanotubes and gold nanoparticles(Au NPs) in a chitosan(CS) matrix. The graphene and the single wall carbon nanotubeswere first placed on the GCE, and the Au NPs were then electrodeposited on the surface by cyclic voltammetry. This structure results in a comparably large surface for immobilization of the capturing antibody(Ab1). The modified electrode was used in a standard sandwich-type of immunoassay. The secondary antibody(Ab2) consisted of Au NPs with immobilized Ab2 and modified with biotinylated DNA as signal tags. Finally, alkaline phosphatase was bound to the biotinylated DNA-Au NPs-Ab2 conjugate via streptavidin chemistry. The enzyme catalyzes the hydrolysis of the α-naphthyl phosphate to form α-naphthol which is highly electroactive at an operating voltage as low as 180 m V(vs.Ag/Ag Cl).Under the optimal conditions, The resulting immunoassay exhibits wide linear range(50 to 3200 pg m L-1)and low detection limit(47pg m L-1)(S/N=3). And it has good specificity and reproducibility. The strategy has a great potential for the sensitive determination of small molecular substances other than prolactin.