| Malachite green with high toxicity and high persistent can residues in aquatic products and environment for a long time and has teratogenicity, carcinogenicity and mutagenicity. The MG residues in aquatic products are clearly limited in the USA and the EU. Malachite green was banned in the veterinary drug list and prohibited to use in food producing animals.At present, the content of MG are usually detected by instrument, but these methods need complex pre-treatment, long analysis time, high cost. However, gold immunochromatographic assay has advantages of easy and simple to handle, taking a short time, low cost. It is suitable for on-site testing and mass screening. We choose MG as a study object to establish GICA strip and use strips to detect the MG in aquatic products.Different sizes of the colloidal gold particle were prepared by reducing the gold chloride with sodium citrate heated by magnetic stirrers. The colloidal gold particle of about 26 nm was suitable for labeling to an anti-MG monoclonal antibody. The optimum p H of solution for labeling was adjusted by using 0.2 mol/L K2CO3 for about 4 μL, the optimum spiking rate of monoclonal antibody was 15 μg/m L, and the optimum spiking rate of 10% BSA was 30 μL/m L.GICA materials chose the sample pad(GL-b01), the Jinbiao pad(GL0194), the absorb pad(H5076) and nitrocellulose(NC) membrane 135 of Millipore. The solution of gold-labelled antibody was diluted four times. The MG-BSA conjugate 2.5 mg/m L was dispensed onto the NC membrane to serve as test line, and sheep anti-rabbit Ig G 0.1 mg/m L served as control line. the sensitivity of strips by the colorimetric method was 10 ng/m L.The strip has high stability, good specificity, without cross-reaction with the leucomalachite green, Chloramphenicol, methylene blue, tetracycline and cross-reaction rate with crystal was66.7%. The results of the application of strips for testing the real negative samples of aquatic product were 0% of false positive rate, coherence with the different standard substance results of LC-MS/MS. When the standard substance of MG was 50 ng/g, the result of strips was 0% of false negative. The recovery rate of the mathod was 60%~80%, and when the concerntration of malachite green was more than 25 ng/g in practical samples, the strip can be accurately datermined.The result of test strip is stable, easy to be justified and suitable for detecting the sulfonamides residues in aquatic products on-site. The method of producing the one step strip for MG could be used in the development of strips for the detection of antibiotic, veterinary drug,etc. |
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