OBTAIN THE COMPONENTS FOR T-2 TOXIN DETECTION IN FEED USING ELISA
The method for prodaction of components for the T-2 definition toxin in food is developed. Cases of fusariotoxicosu that caused by T-2 toxin and produced by F. Sporotrichioides fungi, are widespread in all areas of Ukraine. Actuality. There are no domestic express methods of diagnosis of toxins in feed and livestock products. The purpose of the work. To develop a methodology of the production of components for T-2 toxin detection in feeds using ELISA. Materials and methods. When performing research we used a dry standard of T-2 toxin, acetonitrile, 10 % of glutarovy aldehyde, a saturated solution of ammonium sulfate, borate buffer, dimethylsulfoxide, Sephadex G-10, DEAE-Sephadex A-50, periodates sodium, acetic buffer, and chromatographic columns. For immunization we used the male rabbits of chinchilla breed. The study was carried out using of immunological, immunochemical, immunoassay, biochemical and physical methods. The results of the research. With the purpose of T-2 toxin definition of in feed it was developed to methodizes for production components ELISA. First, it was necessary to conduct the conjugation mycotoxin with BSA. Future there, were applied two schemes of antigen conjugation, namely with dimethylsulfoxide (1) and without the addition of dimethyl sulfoxide (2). It have been added 10 % glutaric aldehyde to antigens obtained by two schemes. In the next stade two conjugates were separated by column chromatography. On the results of experiment antigen conjugation in borate buffer solution with the adding of dimethylsulfoxide has been more active. After studying the physico-chemical properties of the conjugates it was conducted rabbits immunization. Blood samples were collected in 10 days after the last immunization. In the future it immunoglobulin G was obtained, using of ion-exchange chromatography from rabbit antisera. Conjugation of received antibodies with horseradish peroxidase was carried using out periodates method by Nakane. Thus, techniques have been developed for the production of components producing ELISA. There were tested various buffer systems, two Chromogen, the optimum titer of the ELISA receiving was defined conjugate. The results ELISA testified that the best results are obtained when using conjugate in 1:500 dilution. Chromogen TMB and buffer CBB. The conclusions. Method for receiving to detect T-2 toxin in food was developed.