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本研究旨在对Doucet 等报道的定量检测大鼠单根近端肾小管Na+-K+-ATPase 活性方法进行改进。取经过Ⅱ型胶原酶消化的大鼠肾脏皮质组织,在体视显微镜下手工分离单根近端肾小管,并测量其长度,经低渗和冻融处理后与[γ-32P]ATP 共同孵育,液闪法检测从[γ-32P]ATP 解离出的32Pi,采用修正后的公式计算Na+-K+-ATPase 活性。改良法与 Doucet 等的方法比较,测定单根近端肾小管 Na+-K+-ATPase 活性无显著性差异(P>0.05)。改进后的方法节省试剂,操作简便、省时。
The purpose of this study was to improve the method of quantitative detection of Na + -K + -ATPase activity in single proximal tubule of rats reported by Doucet et al. Rat renal cortical tissue digested with type II collagenase was isolated and manually isolated from the proximal proximal tubule by stereomicroscope. The length of the tubular was measured and incubated with [γ-32P] ATP after hypotonic and freeze-thaw treatment , 32Pi dissociated from [γ-32P] ATP was detected by liquid-flash method and the Na + -K + -ATPase activity was calculated using the corrected formula. Compared with the method of Doucet et al by modified method, there was no significant difference in the activity of Na + -K + -ATPase in single proximal tubule (P> 0.05). The improved method saves reagents, easy to operate and saves time.