台灣飯匙倩心臟毒熒光衍生物的製備與性質

Abstract

心臟毒素以異硫氰酸熒光素標誌成?熒光毒素混合物之後，以Sephadex G－25及CM Cellulose C－52柱形層析法製備及純化，可以得到二個不同標誌程度的熒光衍生物。以Ⅰ及Ⅱ代表其一些性質，比較如下： FTH-CTX(Ⅰ) FTH-CTX(Ⅱ) NATIVE CTX 接以FITC程度〔FITC〕／〔CTX〕 1.45 0.99 電泳移動快慢 最慢 次之 最快 溶血作用活性（以六小時?準） 50.9％ 71.5％ 90.1％ 免疫活性 ＋ ＋ ＋ 熒光強度 1.23 0.61

Taiwan Cobra Cardiotoxin (CTX) was reacted with near stoichiometric quantities of Fluorescein-isothiocyanate (FITC) in 0.1 H. borate buffer at room temperature for 15 min. to yield the correspoding N-fluoresceinylthiocarbamoyl CTX. The reaction products were characterized by biological, biochemical, and spectroscopic methods. These were summarized below. 1. Spectrophotometric analysis at 495 mu and protein content measurement indicated that a monofluorescein CTX was separated from unconjugated CTX and other reaction products by ion exchange chromatography. (degree of substitution I 0.99 and II 1.45, respectively) 2. Dye-toxin conjugates had the same hemolytic properties as native toxin, but their potencies were lower. (activity, 71.6% and 50.9%, respectively) 3. Like a pronounced decrease in hemolytic activity was observed, the integrity of antigenic specificity also altered. But the cross-reaction with anti-native CTX antibody to Dye-toxins both remains. 4. The emission maximum of the fluorescent cardiotoxin shifted, from 530 to 519 mu. This suggested a hydrophobic environment of the FITC binding sites. 5. When excited at 280 nm, the fluorescence of the dye bound increases dromatically but the fluorescence of conjugated CTX decrease. According to Forster, resonance transfer of energy could take place between the dye and the protein tyrosine.