The Research On Visual Detection Of Salmonella Based On HCR

Food poisoning increased year by year,foodborne pathogenic bacteria has been one of the most crucial factors,which affect the food safety of our country extremely.Salmonella is one of the most important foodborne pathogenic bacteria,it exists in the vegetables,fruit and milk without disinfection and sterilization generally.Its host is human and animals,which could cause livestock and fowl death extensively and financial loss.And it exists in the products of animals as infection source,which threatens the health and safety of human beings and animals.However,normal detection methods exist with a series of problems as complicated operation,time-consume,large-scale operating instruments,costs of antibody and enzyme,the poor sensitivity and unstable specificity.So in order to improve the shorts of normal methods,it is significant for us to detect Salmonella in food sensitively and promptly.To achieve the quick and low-cost detection,two detect methods of Salmonella was set up.First,the method is that Capture Probe(CP)modified with biotin and Detect Probe(DP)with FITC was designed to match with target Salmonella sequence according to the principle of complementary base pairing.The former can be captured by streptavidin(SA)immobilized on microwell plate and the later can conjunct to anti-FITC-HRP.When target is present,sandwich “CP/target/DP” was formed and HRP was also immobilized on the well.The quantity of Salmonella sequence can be estimated by observing HRP catalyzing TMB turning from colorless into blue.The experiment was consisted of three parts: optimal conditions,sensitivity and specificity.The result showed that when the reaction time of CP,anti-FITC-HRP and FITC are both 2 h,the performance is best.The detect limit of this method is 22.91 pmol/L and it can discriminate some other foodborne pathogenic bacteria with target,showing high sensitivity and good specificity.It is suitable for multiple detection once,with good applicability and the whole procedure can be completed in 6 hours.In second study,coupling with multiple HCR(hybridization chain reaction)concatemers,16 S rRNA of food-borne Salmonella was functionalized in a signal structure for lateral flow nucleic acid biosensor(LFNAB)detection.16 S rRNA was incubated with two specific capture probes and a multitude of helper probes carrying the same initiator,to unwind its secondary structure to form “initiators-on-a-string” complex.By means of initiators,each target 16 S rRNA yielded multiple HCR concatemers tethered numerous biotins,and introduced numerous Streptavidin-labeled gold nanoparticles on the LFNAB.The reaction time is 30 min.The limit of detection is 53.65 CFU/mL for Salmonella.Significantly,this method has high specificity and applicability in the detection of Salmonella in food samples.In conclusion,this visual detection system is a feasible method for non-specialist personnel and point-of-care diagnosis in low-resource settings.