tailieunhanh - Báo cáo khoa học: "Development of a novel antigen capture-ELISA using IgY against porcine interleukin-6 and its application"

Tuyển tập các báo cáo nghiên cứu khoa học quốc tế về bệnh thú y đề tài: Development of a novel antigen capture-ELISA using IgY against porcine interleukin-6 and its application | J. Vet. Sci. 2004 5 4 337-343 JOURNAL OF Veterinary Science Development of a novel antigen capture-ELISA using IgY against porcine interleukin-6 and its application Deog Yong Lee Young Wook Cho Sang Gyun Kang Sung Jae Shin Han Sang Yoo Department of Infectious Diseases College of Veterinary Medicine and School of Agricultural Biotechnology Seoul National University Seoul 151-742 Korea Interleukin-6 IL-6 is introduced as a marker of disease. At present a variety of method may be used to quantify expression of this protein. Antigen capture-ELISA is a sensitive and accurate quantification method previously used with ovine rat and human IL-6 proteins. However it has never been reported to quantify porcine IL-6 protein using capture ELISA. In this study we generated and characterized a set of IgY and monospecific polyclonal antibodies to recombinant porcine IL-6 rpIL-6 and combining these with a sensitive and specific capture-ELISA for a diagnostic purpose. cDNA encoding the mature protein coding region of porcine IL-6 was cloned and expressed with pQE-30UA expression vector. rpIL-6 was then expressed and purified by using Ni-NTA resin. Protein mass of 24 kDa was found with SDS-PAGE and the identity of the protein was confirmed by Westernblot. Production of polyclonal antibodies against rpIL-6 was performed using the purified rpIL-6 in mice and hens. An antigen capture-ELISA was developed with the antibodies after their extraction. To compare the IL-6 level in the different sanitary state of farms pig sera were randomly collected and concentration of IL-6 in the sera was measured with the antigen capture-ELISA. The capture-ELISA with the optimal concentration of antibodies in this study was able to detect about 10 ng ml of rpIL-6. IL-6 levels determined with the capture-ELISA in pig sera showed positive correlation with the sanitary states of the farms. These results suggested that the developed antigen capture-ELISA could be a good tool for the screening of microbial .