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Rac1磁珠激活下拉(Pull-Down)试剂盒
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Rac1磁珠激活下拉(Pull-Down)试剂盒

Rac1磁珠激活下拉(Pull-Down)试剂盒,Rac1 Pull-Down Activation Assay Biochem Kit (Bead Pull-Down Format),非常适合开始WB和SDS-PAGE实验类型。更多视频请关注视频号【艾维缔】。哔哩哔哩【IVDSHOW】。抖音【军哥聊表观】。
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Rho 开关通过在活跃的 GTP 结合态和不活跃的 GDP 结合态之间交替运行。 了解调控 GTP 酶激活/失活的机制显然具有重要的生物学意义,也是目前正在深入研究的课题。 许多 Rho 家族效应蛋白会特异性识别蛋白的 GTP 结合形式,这一事实已被实验利用,开发出一种强大的亲和纯化检测方法,用于监测 Rac 和 Cdc42 蛋白的活化。 该检测方法使用了 Cdc42 / Rac 效应蛋白 p21 活化激酶 1(PAK)的 Cdc42/Rac 交互结合(CRIB)区域(也称为 p21 结合域,PBD)。 事实证明,CRIB/PBD 蛋白基团能与 Rac 和/或 Cdc42 蛋白的 GTP 结合形式特异性结合。 事实上,PAK 的 PBD 区域对 GTP-Rac 和 GTP-Cdc42 都有很高的亲和力,而且 PAK 的结合会导致 Rac 和 Cdc42 的内在水解率和催化水解率显著降低,因此它是亲和纯化细胞裂解液中 GTP-Rac 和 GTP-Cdc42 的理想工具。 本试剂盒中的 PAK-PBD 蛋白对应于 67-150 个残基。 其中包括高度保守的 CRIB 区域(aa 74-88)以及与 GTP-Rac 和 GTP-Cdc42 高亲和力相互作用所需的序列。 PAK-PBD 以 GST 融合蛋白的形式存在,可以用谷胱甘肽亲和珠 "拉取 "PAK-PBD/GTP-Rac(或 GTP-Cdc42)复合物。 因此,该检测提供了一种量化细胞中 Rac 或 Cdc42 活化的简单方法。 活化的 Rac 量可通过使用 Rac 特异性抗体进行 Western 印迹来确定。

Rac1磁珠激活下拉(Pull-Down)试剂盒具有如下优势和特点:

  • 分析体内 Rac1 激活水平。
  • 检测能增强 Rac1 活性的化合物和蛋白质。
  • 检测抑制 Rac1 活性的化合物和蛋白质

图 1. Cdc42磁珠激活Pull-Down试剂盒中的谷胱甘肽琼脂糖珠颜色鲜艳,使试剂盒易于使用。

图 2.通过用 GTPγS 或 GDP 加载细胞裂解液中的 Rac1 蛋白,对 Rac1 活化试验进行了测试。不出所料,GTPγS 负载的 Rac1 非常有效地沉淀下来,而 GDP 负载的 Rac1 则很少沉淀下来。

图 2. BK035-S Rac1 活化检测的结果。使用试剂盒 BK035-S 在 Western 印迹中沉淀并检测活化的 Rac1。第一条泳道显示的是 50 ng 重组 His 标记的 Rac1 标准品(重组 His-Rac1)。随后的泳道显示从等量细胞裂解液中拉取无活性的 GDP 负载 Rac1(Rac1-GDP PD)或活性的 GTPγS 负载 Rac1(Rac1-GTP PD)。

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Rac1磁珠激活下拉(Pull-Down)试剂盒 CNBK035-01 20次 2-8°C
Rac1磁珠激活下拉(Pull-Down)试剂盒 CNBK035-02 50次 2-8°C

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