Methodology Research Of Screening SsDNA Aptamers Of Oxoplasma Antibody And Cytomegalovirus Antibody Using ELEX And Use SPR Technology For Diagnosis

Backgroud:Birth defects to family, country and society brings great mental and economic burden. Atpresent, clinical on fetal congenital defects no effective therapeutic method. Birth defectsprevention method, early pregnancy prevention is most effective. TORCH is a group ofpregnant mice infected with pathogenic microorganisms, can pass through the placenta orbirth canal caused by intrauterine infection, leading to miscarriage, stillbirth or fetal growthretardation, congenital malformations, neonatal infection and infant, child growth anddevelopment disorders. Our country each year about26000children with TORCH infectionwas born, average each hour3, therefore, the TORCH screening is pregnant, infectious diseaseis the important content of. However, according to statistics, China’s current diagnosticclinical commonly used ELISA method, the gold standard method of testing of pregnantwomen age on serum TORCH-IgG and TORCH-IgG levels, the false positive rate is higher,can provide accurate diagnosis on the basis of clinical.Objective:To screen oligonucleotide aptamers specific binding to Toxoplasma antibody andCytomegalovirus antibody using Systematic evolution of ligands by exponential enrichment(SELEX). To establish a rapid detection technique is based on aptamer-based surface plasmonresonance (SPR) microarray by the TOX and CMV IgG antibody detection to realized the realtime and on line the detection of the microbial.Methods:Synthesize an ssDNA library of random78nucleotides in vitro and use SELEX to screenthe library. Screen for12rounds using the Toxoplasma protein antibody and Cytomegalovirusantibody as the target material and detect the binding of oligonucleotides and protein usingthe biotin-avidin system. Clone and sequence the library of screened aptamers, then use thesoftware DNAMAN to analyze the structure and do primary test of the acquired aptamers by sandwich assay. The sandwich assay is composed of aptamers which can capture and detect.Results:We use the sandwich assay to detect75serum samples. With the cut-off value being1.42,the negative percentage was84.6％(Toxoplasma)and92.3%(cytomegalovirus) in the negativegroup and the positive percentage was87.1%(Toxoplasma)and90.3%(cytomegalovirus) in thepositive group, which showed certain detective worth. It showed that the detection methodhave good precision and detection range or could realize on line the detection of targetsequence. One of the system could detect from20umol/L to300umol/L.Conclusions:A set of ssDNA aptamers with considerable binding affinity with Toxoplasma antibodyand Cytomegalovirus antibody were successfully selected from the initial random DNAlibrary and some certain detective worth was showed and there was potential for detectingClinical specimens...