| Nitrofurans drug is a kind of synthetic with 5-nitrofuran ring mother nucleus of antibacterial drugs, mainly includes nitrofurazone, nitrofurantoin, nitrovin, furazolidone, furaltadone, nifurstyrenate sodium, nifursol, nifuroxazide, etc.The drugs,which are broad antibacterial spectrum, high efficiency and low price, arewidely used in livestock, poultry and aquatic animal disease prevention and treatment. According to the previous study, the nitrofurans drug for livestock and poultry has toxicity, and is also a kind of potentially carcinogenic and mutagenic substances.Nitrofurans drug residues in animal food have potential health riskathuman.Therefore, Nitrofurans ban the use of world.At present, the detection method of nitrofurans drug mainly HPLC, LC-MS, LC/MS/MS and ELISA, and the detection methods are mainly nitrofurans metabolites in animal food.There are few literatures reported in detection of nitrofurans drug technical in feed. The high precisiondetection methods used nowrequire specialized equipment, and the test time is long, not easy to popularize. However, the colloidal gold strip is based on the antigen antibody specificity, hashigh sensitivityand the test time is fast,and don’t need professional operation,and so on.So the colloidal gold striphas a broad application prospect.The aim of this study is to develop a kind of detection,which can detect the nitrofurans drug in livestock and poultry feed, andcompletely eliminate nitrofurans drug harm to human body.In this study,we used 5-nitrofuran-2-formaldehyde astemplate,then synthesiednitrofurans drug of haptensusing the methods of CDI and NHS respectively.We immuned10 Balb/c mouseused those two immunogen respectively.After the immune blood testing, we select the high titerserumto preparation of single cell fusion of the spleen. After purification of ascites titer of1:1.8×106. Nitrofurazone IC50 was0.339ppb, furazolidone IC50 was0.462ppb, nitrofurantoin IC50 was0.304ppb, furaltadone IC50 was0.535ppb, nifurstyrenate sodium IC50 was0.486ppb, nifursol IC50 was0.578ppb, and nitrovin IC50 was0.578ppb.The particle sizes prepared by sodium citrate reduction method were20 nm,30 nm,40 nm colloidal gold particles, we finally chose to use 30 nm size of colloidal gold, after doing a series of colloidal gold particles selectionexperiments. Through the screening of NC membrane, gold standard pad, sample pad, blotting paper and paired antibody, we determined the best materials. We developed the specificity and sensitivity experiments, and this product had nitrofurans drug reaction specificity, did not have specificity reactionwith BSA, clenbuterol and other drugs. The detection limit of nitrofurazone, furazolidone, nitrofurantoin, furaltadone, nifurstyrenate sodium, nifursol and nitrovin was30 ng/ml,40 ng/ml,30 ng/ml,50 ng/ml,40 ng/ml,50 ng/ml and 80 ng/mlrespectively. We did the experiments of processingsamplesby direct dilution and feed add recycling experiment, and two detection methodsby ELISA and HPLC.which validated the effectiveness of the strip. Thebetween batch and batch repeatability is good, and through 3 months aging tests, results shown that the strip can be successfully used in the feed nitrofurans antibiotics detection.The strip can be analyzed without instruments, and the operation is simple, rapid,and the results can be read in 10 minutes. The experimental results have high stability, and the strip can be used to do nitrofurans drug scene of livestock and poultry feed when a large number of screening effective means, also be used for other agricultural development of veterinary drug residue detection strip for reference. |
