不同方法测定血红蛋白溶液中总蛋白量对比分析.docVIP

不同方法测定血红蛋白溶液中总蛋白量对比分析 　　摘要:目的 采用三种方法对含有大量血红蛋白溶液的样本进行总蛋白量的测定，并进行对比，确定一种可行且简单的测定血红蛋白溶液中总蛋白量的方法。方法 分别采用BCA法、Bradford法和Lowry法测定溶液总蛋白量，并采用HiCN法测定溶液中血红蛋白的含量。比较不同方法测定数据的差异。结果 5份样本用BCA测得总蛋白量为76.33g/L，Bradford法测得值为179.59g/L，Lowry法测得值为22.81g/L，HiCN法测得血红蛋白量为133.65g/L。结论 Bradford法是一种可行且重复性好的测定血红蛋白溶液中总蛋白量的方法，BCA法和Lowry法的测定值明显低于实际值。 ? 　　关键词 血红蛋白溶液 总蛋白量?? 　　 　　The Compsrison of Different Handwork Methods on Measuing the Total Protein Concentration of the Hemoglobin Solution.Cai Ming-cui.The Blood Center of Liangshan,Xichang 615000,Sichuan,China.?? 　　【Abstract】Objective Three methods commonly used do determine the protein concentration were taken to measure the total protein of the solution containing stroma-free hemoglobin. And we tried to confirm a feasible way on measuring the total protein concentration through comparison of different methods.Methods The BCA, Bradford and Lowry methods were used to determine the total peotein, and the HiCN method was exploited to measure the level of hemoglobin.Results The average concentration of the total protein of 5 samples was 76.33g/L by BCA,179.59g/L by Bradford, and 22.81g/L by Lowry. The average concentration of hemoglobin by HiCN method was 133.65g/L.Conclusion The Bradford method was feasible and precise to determine the total protein concentration of hemoglobin solution. The results of BCA and Lowry methods were obviously lower than that of Bradford method and even the level of hemoglobin.? 　　【Key words】Hemoglobin solution;Total protein concentraton?? 　　 　　自Amberson等［1］首次使用粗制血红蛋白溶液作为血液代替品以来,研究人员通过层析［2］、超滤［3］、制备型HPLC等［3］方法纯化得到了高纯度的血红蛋白溶液，并以此为基础进行各种修饰，开发出不同的红细胞代用品［4］。在此过程中，血红蛋白纯度的检测是一个关键指标，影响化学修饰和产品的化学特性。文献报道多采用HPLC发，但该方法复杂，且仪器昂贵。笔者试图通过对常用测定蛋白质含量的方法（BCA法、Bradford法、Lowry法）的测定结果比较，为制备血红蛋白溶液寻找一种简单、快速的检测方法。? 　　 　　1 材料与方法? 　　 　　1.1 血红蛋白溶液样品的制备 新鲜采集的血样按文献［5］方法制备并编号。? 　　1.2 血红蛋白含量测定 对上述制备的5份样本按HiCN［5］测定，并应用MD2型库尔特计数仪检测。? 　　1.3 总蛋白测定 取40ul血红蛋白溶液加入10ml容量瓶中，稀释成1/250浓度，进行下列测定（每份样品均同）。 样品总蛋白量=下述各方法所得蛋白量×稀释倍数（250）? 　　1.3.1BCA法 取4个E