Basic Study On Detection Of Serum Circulating Antigen By Gold Nanorods With Specific IgG Antibody Against Clonorchis Sinensis

Objective To prepare high purity specific IgG antibody against hepatic trematode and optimize the most appropriate concentration of specific IgG antibody labeled on the surface of nano-gold rods,obtain functional nano-gold rods with specific IgG antibodies against hepatic trematode and apply them to detect serum circulating antigen.To study the detection effect of functional nano-gold rods on serum circulatingantigeninearlyorlowinfectionofhepatictrematode.Method1.Seed-mediated growth polymerization of stable gold nanorods with different diameter-length ratios.2.Collecting the antiserum of Clonorchiasis hepatica by establishing animal models of clonorchiasis hepatica.The specific IgG antibody of hepatic trematode was purified by column chromatography and the purified effect was identified by SDS-PAGE.The sulfhydrylated specific IgG antibody was combined with gold nanorods.The red shift of longitudinal plasma resonance absorption peak on the surface of gold nanorods before and after scanning labeling by ultraviolet spectrophotometer was compared,and the most appropriate coating concentration was selected to constructfunctionalizedgoldnanorods.3.Screening serum circulating antigen related components.The antigen bands of excretion and secretion of hepatic trematode at each stage were purified by SDS-PAGE and eluted to collection.The excretory and secretory purified antigens of each stage were mercaptoylated and combined with nano-gold rods to construct functional nano-gold rods for detection of positive serum antibodies against hepatic trematode.To observe the red shift of plasma resonance absorption peak on the surface of gold nanorods before and after ultraviolet spectrophotometer scanning,analyze the immune response of excretory and secretory antigens,and identify the antigenicity of circulating antigensinserum.4.Serum circulating antigens of different infection time,different infection degree and different dilution degree were detected by nanogold rods functionalized with specific IgG antibody of hepatic trematode,compared with the ELISA results of rat hepatic trematode,the detection advantages of functionalizednanogoldrodswereanalyzed.Result1.Established successfully of rat model of hepatic trematode cysticercosis infection and collection of different infection degrees and different infection time serum circulating antigens.High purity of specific IgG antibody against hepatictrematodewas obtained.2.Ultraviolet spectrophotometry was used to detect specific IgG antibody against hepatic trematode at 20μg/m L concentration labeled with gold nanorods.The maximum red shift of surface longitudinal plasma resonance absorption peak was 20 nm compared with that before labeling,and the peak shape was normal.The optimum coating concentration was 20μg/m L.3.Screened serum specific excretion-secretion purified antigens with strong antigenicity and determined serum circulating antigens as serum diagnostic antigen.4.Functionalized nanogold rods with specific IgG antibodies against hepatic trematode could be detected on the 3rd day of mild to moderate to severe infection.Serum circulating antigens of 50 cysticerci could be identified by positive results.ELISA was positive on the 17 th day of mild infection and on the 10 th day of moderate and severe infection.The positive time of functionalized gold nanorods was 14 days and7 days earlier than that of ELISA.Functionalized nanogold rods could detect positive serum circulating antigens with a maximum dilution of 1:400,and the ELISA assay could detect a maximum dilution of 1:100.ConclusionGold nanorods with adjustable aspect ratio were prepared by optimized seed-mediated method.The optimum concentration of specific IgG antibody labeled on gold nanorods was 20μg/m L.To obtain the detection results of functional nanogold rods with specific IgG antibodies against hepatic trematode.Compared with ELISA results,functional gold nanorods have the advantages of sensitivity and early detection,simple operation and low cost,which provide new technical support for the early diagnosis of hepatic schistosomiasis.