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为了探索在中性环境中P—NPP酶活性的检出方法,对作为捕捉剂的铅盐种类进行了对比和选择。实验动物为体重35—40g的ddy系小白鼠。方法:用含20％多聚甲醛和0.5％戊二醛的固定液灌流固定10分钟,取出肾脏,做20—40μm厚的未冻切片,分别投入下面五组反应液中,37℃,浸渍30分钟。反应液的组成为:Tricine-NaOH缓冲液2.5mM,pH7.4;p-NPP10mμ;KC150mμ;levamisole2.5mμ;DMSD25％;再各个加入柠檬酸铅、硝酸铅、硫酸铅、氯化铅、醋酸铅4mμ。反应后,光镜用1％硫化铵液浸1—2分钟,电镜则用1％OsO_4后固定。再进行常规处理、分别用光镜和透视电镜观察、照像。
In order to explore the detection method of P-NPPase activity in neutral environment, the comparison and selection of Pb salt species as capture agent were carried out. Experimental animals for the weight 35-40g ddy Department of mice. Methods: Perfusion fixed with 20% paraformaldehyde and 0.5% glutaraldehyde for 10 minutes. The kidneys were taken out and unfrozen sections of 20-40 μm thickness were made. The cells were respectively put into the following five reaction mixtures and immersed at 37 ℃ for 30 minute. The composition of the reaction solution was: Tricine-NaOH buffer 2.5mM, pH 7.4; p-NPP 10mμ; KC 150mμ; levamisole 2.5mμ; DMSD 25%; and further adding lead citrate, lead nitrate, lead sulfate, lead chloride, . After the reaction, light microscopy with 1% ammonium sulfide immersion 1-2 minutes, electron microscopy was fixed with 1% OsO_4. Then the conventional treatment, respectively, with light and TEM observation, photography.