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題名 | 雌二醇酵素免疫分析法之建立= |
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作者 | 陳婷婷; 毛嘉洪; |
期刊 | 臺灣畜牧獸醫學會會報 |
出版日期 | 19931200 |
卷期 | 62 1993.12[民82.12] |
頁次 | 頁65-76 |
分類號 | 437.245 |
語文 | chi |
關鍵詞 | 雌二醇酵素色疫分析法; |
中文摘要 | 本研究目的在建立-安全且簡易的酵素免疫分析法(ELISA),以分析樣品中雌二 醇(estradiol, E2)濃度。我們以雌二醇-6-血清蛋白連結體(E2-6-BSA)為免疫原,免疫四 隻兔子以收集抗體;再用蕖草根過氧化酵素(horseradish peroxidase)合成的標幟連結體 E2-3-HRP與E2-6-HRP進行固相競爭型酵素免疫分析。結果顯示,於吸附萬分之一具高靈敏 度抗體的微滴盤中加入樣品與八萬之一的標幟連結體競爭,可得最佳分析結果。當以 E2-3-HRP分析時,四隻兔子的抗體可分別測到13.2、52.0、10.3及6.3pg/50 μ1的雌二醇; 若以E3-6-HRP分析,則可達到61.7、328.3、3.6與1.3pg/50μ1的靈敏度。進行交叉反應 時,除雌素酮(estrone)及去氫異構男性脂酮(dehydroisoandrosterone)具有1.5%和0.001 %的結合率外其餘均小於0.00001%,顯示此抗體對雌二醇具有高度特異性。注射孕馬血清 30日齡小鼠,將其卵巢均質化,並以NADPH和男烯二酮(androstenedione)培養,可見雌二 醇量隨培養時間的加長而增加。又皮下給予小鼠不同劑量孕馬血清,取完整卵巢於DME/F-12 溶液中培養4小時,雌二醇分泌量亦隨給予劑量而有所差異。經以上結果顯示此酵素免疫分 析法可應用於樣品中雌二醇濃度之偵測。 |
英文摘要 | A simple and safe enzyme-linked immunosorbent assay (ELISA) of 17β -estradiol was established in this study. Four California white rabbits were immunized with bovine serum albumin conjugated with estradiol at 6 position. Estradiol conjugated at 3 or 6 position with horse-radish peroxidase (HPR) were used in the solid phase competitive reaction. The best titer for antiserum and enzyme conjugate were 1/10,000 and 1/80,000, respectively. The detectability for each four rabbit's antiserum was 13.2, 52.0, 10.3, and 6.3 pg/well, respectively, when using estradiol-3-HRP as conjugate. When using estradiol-6-HRP as conjugate, the detectability was 61.7, 328.3, 3.6, and 1.3 pg/well, respectively. The crossreactivity with estrone was 1.5% and with dehydroisoandrosterone Was 0.001%. The crossreactivity with other sex steroid hormones and phenol red were less than 0.00001%. Ovarian tissue homogenates from pregnant mare serum gonadotropin (PMSG) pretreated mice cultured with NADPH and androstenedione showed a linear increase of estradiol synthesis from 0 to 40 mimutes. Mice injected with different level of PMSG also showed an increase of estradiol secretion from intake ovary culture in Dme/F12 media. This estradiol ELISA is satisfied in estradiol analysis. |
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