Cardinaud R, Dassin E, Bourebia J
Thromb Haemost. 1983 Dec 30;50(4):824-30.
Synthesis of actin is a reasonably correct representation of the platelet production rate. Measurement of actin production obviates some of the drawbacks encountered in currently available methods. Platelet actin was characterized on polyacrylamide gel electrophoreses using methylated radioactive rabbit actin as a reference. Platelet actin was unambiguously identified by showing that it forms a complex with DNAase-I similar to the complex obtained with pure rabbit actin. Six days after intravenous injection of 75Se-seleno-methionine actin was one of the most highly labelled platelet components. The platelet pellet (one rat per sample) was solubilized with 1% Triton X-100, 0.75 M guanidine-HCl and dialyzed in a medium containing 1% SDS (to eliminate Triton X-100 and guanidine-HCl). A carefully measured aliquot was deposited on a polyacrylamide gel. After electrophoresis in the presence of SDS the radioactivity of the actin band was measured and the ratio of the total actin radioactivity to the injected radioactivity was taken as a measure of platelet production. The validity of the actin probe was tested with populations of normal animals. The specific radioactivity of actin was proportional to the injected dose of 75Se-seleno-methionine up to 1 mCi/kg animal. A semi-log plot of actin specific radioactivity vs time exhibited a pseudo-first order decrease. Another constituent with a high specific radioactivity (XM) was excluded as a suitable probe because it was shown to be an adsorbed plasma protein.
肌动蛋白的合成是血小板生成速率的一种合理准确的体现。肌动蛋白生成量的测定避免了现有方法中存在的一些缺陷。在聚丙烯酰胺凝胶电泳中，以甲基化放射性兔肌动蛋白作为参照对血小板肌动蛋白进行了表征。通过表明血小板肌动蛋白与脱氧核糖核酸酶 -I 形成的复合物类似于用纯兔肌动蛋白得到的复合物，明确鉴定了血小板肌动蛋白。静脉注射75Se - 硒代蛋氨酸6天后，肌动蛋白是血小板中标记程度最高的成分之一。将血小板沉淀（每个样品一只大鼠）用1% Triton X - 100、0.75 M 盐酸胍溶解，并在含有1% SDS的介质中透析（以去除 Triton X - 100 和盐酸胍）。将仔细测量的一份样品加样到聚丙烯酰胺凝胶上。在SDS存在下进行电泳后，测量肌动蛋白条带的放射性，并将总肌动蛋白放射性与注射放射性的比值作为血小板生成的指标。用正常动物群体对肌动蛋白探针的有效性进行了测试。肌动蛋白的比放射性与75Se - 硒代蛋氨酸的注射剂量成正比，直至1 mCi/kg动物。肌动蛋白比放射性对时间的半对数图呈现出拟一级下降。另一种具有高比放射性的成分（XM）被排除作为合适的探针，因为它被证明是一种吸附的血浆蛋白。
