Detection of Antibody against Avian Leukosis Virus Subgroup a by Enzyme-linked Immunosorbent Assay with Baculovirus Expressed gp85 Protein

利用桿狀病毒表現家禽白血病A亞群gp85蛋白質以酵素連結免疫吸附法檢測抗體

許萌芳(Meng-Fang Hsu) ； 王金和(Ching-Ho Wang) ； 萬灼華(Cho-Hua Wan)

《臺灣獸醫學雜誌》 40卷1期 (2014/03) Pp. 39-45

摘要

為了檢測家禽白血病A亞群病毒，（avian leukosis virus subgroup A，ALV-A）的抗體，選殖台灣分離ALV-A的gp85基因，利用桿狀病毒載體表現系統進行ALV-A之表面醣蛋白gp85表現。重組桿狀病毒感染後之昆蟲細胞，進行西方墨點法分析及免疫螢光染色法可偵測到重組gp85蛋白質之表現。重組gp85蛋白質直接做為塗鍍抗原，與標準陽性和陰性血清進行ELISA測試，確認ALV-A抗血清可造成ELISA讀值上升，SPF雞隻血清則否。對田間樣本血清進行測試，以血清中和試驗做為抗體檢測為金標準，得到ELISA之敏感性為80%（24/30），特異性為83.3%（35/42）。因此利用桿狀病毒表現系統進行之ALV-A gp85蛋白表現及酵素連結免疫吸附法可應用於現場抗ALV-A抗體的檢測。

關鍵字

抗體的檢測 ； 家禽白血病病毒 ； 桿狀病毒載體 ； 酵素免疫連結吸附法 ； A亞群

並列摘要

To detect antibody against avian leukosis virus subgroup A (ALV-A), the gp85 gene of an ALV-A strain isolated in Taiwan was cloned and expressed in a baculovirus expression vector. The expressed protein confirmed by Western blotting and immunofluorescence tests with anti-ALVA antiserum was coated in 96-well plates and was used as an antigen for antibody detection by an indirect enzyme-linked immunosorbent assay (ELISA). Neutralization test performed in chicken sera from the infected and uninfected flocks was used as the standard to evaluate the accuracy of the ELISA. The cut-off point was calculated by receiver operator characteristic (ROC) curve to be 1. The sensitivity and specificity of this ELISA were 80% (24/30) and 83.3% (35/42), respectively. In conclusion, this ELISA could be applied for the anti-ALV-A antibody detection in the field.

並列關鍵字

antibody detection ； avain leukosis virus ； baculovirus vector ； enzyme-linked immunosorbent assay ； subgroup A