乙酰胆碱酯酶固定化方法的研究

Study on Immobilization Method of Acetylcholinesterase

以戊二醛为交联剂,牛血清白蛋白(BSA)为保护剂,将乙酰胆碱酯酶(AChE)交联固定到商品载体上,制备固定化酶片。对影响酶固定化的重要因素进行了考察,获得了最佳固定化条件。实验结果表明,以孔径为0.45μm的硝酸纤维素滤膜作载体,乙酰胆碱酯酶用量10U,5%(体积分数)戊二醛2μL,1%(质量分数)BSA10μL,配成70μL的酶溶液,3°C固定8h,可获得较好的固定化效果。不同批次制备的酶片,其活力值标准偏差为3.27%～5.03%,酶片在0.1mol LpH8.5磷酸盐缓冲溶液中3°C下可保存60d。

Using glutaraldehyde as a cross-linking agent and bovine serum albumin as a protectant, an immobilization method of acetylcholinesterase is developed. The important factors affecting immobilization are explored, and the optimum conditions are obtained. The best immobilization conditions are:10 U of acetylcholinesterase, 2 μL of 5% glutaraldehyde, 10 μL of 1% bovine serum albumin, 3°C, 8 h, and a cellulose nitrate membrane filter with a pore size of 0.45 μm as the carrier membrane. The activeity of enzyme membranes produced in different batches is quite similar (RSD 3.27%～5.03%), and the enzyme membranes is stable for 60 d in 0.1 mol/L phosphate buffer(pH 8.5) at 3°C.