乳鼠成骨细胞碱性磷酸酶活性检测方法初探

目的探讨乳鼠成骨细胞碱性磷酸酶活性的检测方法,为研究骨代谢异常及骨质疏松症等骨病提供简便可靠的检测手段.方法采用经体外培养的乳鼠成骨细胞,制备上清液、裂解液和冻融液样本,分别用二乙醇胺、2-乙基氨基乙醇、2-氨基-2-甲基-1-丙醇3种不同缓冲液的试剂方法,测定其碱性磷酸酶活性,并作比较分析.结果使用二乙醇胺缓冲液的试剂方法,对3种标本的测定值都最高,与各组比较分析,差异均有显著性(P＜0.01),且变异系数最小,其他两种方法的测定结果无明显差异(P＞0.05).用3种不同缓冲液试剂方法检测3种不同方法制备的标本,均以裂解液中碱性磷酸酶活性最高,冻融液次之,上清液最低,分析结果,差异均有显著性(P＜0.01).结论用二乙醇胺缓冲液的试剂方法,测定体外培养乳鼠成骨细胞裂解液碱性磷酸酶的活性,效果较为理想.

Methodological discussion on determination of osteoblast alkaline phosphatase in sucking rat

[Objective] To study the methodology on the determination of osteoblast alkaline phosphatase in sucking rat in order to provide a simple and reliable means for bone diseases,such as bone metabolic abnormality and osteoporosis etc.[Method] The osteoblast from sucking rat was cultured in vitro to obtain the samples of supernatant,spliting liquid,freezing-melting liquid.The alkaline phosphatase in samples above were determined using the different buffer solutions of diethanol,2-ethylaminothanol and 2-amino-2-methyl-1-propanol and the results were analyzed.[Results] The activities of alkaline phosphatase in three types of samples above and precision were the highest in diethanol buffer solution and had obvious differences as compared to other two groups(P <0.01),but the results of other two methods had no difference(P >0.05).In three types of samples,the results of alkaline phosphatase were different and had statistical diversity by the three methods(P <0.01),the highest in spliting liquid and the lowest in supernatant.[Conclusion] The method using diethanol as buffer solution is more ideal for the determination of osteoblast alkaline phosphatase activity in cultures of sucking rat in vitro.