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目的:利用125I标记血管抑素(angiostatin,AS),并研究标记产物125I-AS的体外稳定性及生物活性。方法:制备的AS经鉴定后,用Ch-T法进行标记,用纸层析法测标记率,用Sephadex-G50柱纯化,产物分别加入牛血清白蛋白(BSA)、生理盐水及不同摩尔比的半胱氨酸以观察其体外稳定性,并通过人脐静脉内皮细胞系ECV304细胞生长抑制试验来检测125I-AS的生物活性。结果:制备的125I-AS标记率可达85%,125I-AS在生理盐水中1、42、4 h的解离率分别为2.3%、5.2%、8.0%,在10 g/L BSA中1、4、24 h的解离率分别为4.6%、9.1%、11.0%。在不同摩尔比半胱氨酸溶液中孵育,125I-AS存在不同程度的解离,摩尔比在5以下时,4 h的解离率不超过11%,表明标记产物在体外较稳定。分别用不同浓度125I-AS和AS作用于ECV304细胞,并以6-氨基己酸作为对照,发现两者对ECV304细胞增殖均有明显的抑制作用,且125I-AS的作用强于单纯的AS,表明125I-AS具有较强的生物活性。结论:用Ch-T法进行125I-AS标记简单易行,125I-AS对ECV304细胞的生长有明显抑制作用,有较强的生物活性,标记产物在体外较稳定。
OBJECTIVE: To use 125I-labeled angiostatin (AS) and to study the in vitro stability and biological activity of the labeled product 125I-AS. Methods: The prepared AS was identified and labeled by Ch-T method. The labeling rate was determined by paper chromatography and purified by Sephadex-G50 column. The products were respectively added with bovine serum albumin (BSA), normal saline and different molar ratios Cysteine to observe its in vitro stability, and the bioactivity of 125I-AS was examined by the human umbilical vein endothelial cell line ECV304 cell growth inhibition assay. Results: The labeling efficiency of 125I-AS reached 85%. The dissociation rates of 125I-AS in physiological saline at 1, 42, and 4 h were 2.3%, 5.2% and 8.0%, respectively. , 4, 24 h off rate was 4.6%, 9.1%, 11.0%. In different molar ratios of cysteine solution incubation, 125I-AS dissociation to varying degrees, the molar ratio of 5 or less, 4 h dissociation rate of not more than 11%, indicating that the labeled product is more stable in vitro. The effect of 125I-AS on the proliferation of ECV304 cells was obviously inhibited by 125I-AS and AS with different concentrations of 125I-AS and 6-aminocaproic acid as controls. 125I-AS showed strong biological activity. CONCLUSION: The 125I-AS labeling by Ch-T method is simple and easy. 125I-AS can significantly inhibit the growth of ECV304 cells and has strong biological activity. The labeled product is stable in vitro.