Pierce c-Myc-Tag Magnetic IP/Co-IP Kit c-Myc-Tag磁性IP / Co-IP试剂盒
货号：88844
规格：40 reactions
价格：6883
产品类型：免疫印迹/组化
品牌：Thermo Fisher
| Thermo Scientific Pierce 磁性 c-Myc 标签 IP/Co-IP 试剂盒包含使用 c-Myc-标签诱饵蛋白进行 c-Myc 融合蛋白免疫沉淀试验或 co-IP 实验的特异性免疫亲和磁珠和试剂。与基于使用蛋白 A/G 微珠进行捕获的传统 IP 程序不同，该试剂盒使用含有预固定化抗 c-Myc 抗体的磁珠。这些 Pierce 抗 c-Myc 磁珠可确保与生物样品中的 c-Myc 标签蛋白复合物发生特异性结合。因为抗体共价连接到微珠上，所以可以轻松地对 IP 靶标进行洗脱和回收，无抗体污染。全套 IP 试剂盒包括磁珠、裂解/洗涤缓冲液、低 pH 值洗脱缓冲液、中和缓冲液、c-Myc 标签阳性对照裂解物和用于 SDS-PAGE 的非还原样品缓冲液。提供手动和自动磁性分离工作流程的方案。提供足够进行 40 次 IP 或 co-IP 试验的组分。源自人 c-Myc 蛋白的 C 端区域的 c-Myc 肽 (EQKLISEEDL) 是用于重组蛋白表达的多种融合蛋白标签之一。Pierce c-Myc 磁性 IP/Co-IP 试剂盒使用特异性、高亲和力固定化抗体（克隆 9E10），用于对细菌和哺乳动物细胞裂解物以及使用 Pierce 人体外翻译试剂盒制备的裂解物中的 c-Myc 标签融合蛋白进行快速免疫沉淀。将微珠与含有 c-Myc 标签蛋白的细胞裂解物一起孵育，捕获融合蛋白，随后对微珠进行洗涤，然后使用低 pH 值洗脱缓冲液或非还原样品缓冲液进行洗脱。实验方案和缓冲液均经优化处理，可最大限度地提高 IP 和 co-IP 反应的效率，可在洗脱样本中富集特定的蛋白交互复合体。抗 c-Myc 抗体可用于通过 Western 印迹分析检测 c-Myc 标签蛋白。 |
| 特点： |
| -特异性磁珠—共价固定化高质量抗 c-Myc 单克隆抗体可实现免疫沉淀产物的高得率-低非特异性结合— 稳定、预封闭的微珠和特异性抗体可尽可能减少脱靶结合-可靠洗脱—低 pH 值洗脱缓冲液可确保 c-Myc-标签蛋白相互作用复合物的回收，无抗体浸出污染-便利快速—全套试剂盒和易于遵循的说明提供了优化方案，可在约 1 小时内进行 IP 或 Co-IP 实验-通用—磁珠与手动和自动磁性分离工作流程（如 Thermo Scientific KingFisher 仪器）兼容 |
| 数据： |
Better immunoprecipitation results with Anti-c-Myc Magnetic BeadsGreen Renillaluciferase c-Myc fusion protein was expressed in 293T cells. For IP,identical aliquots of the cell lysate were incubated in duplicate for one hour at room temperature with anti-c-Myc magnetic beads from each manufacturer. For all conditions, IP products were eluted identically using low-pH buffer. Eluted fractions (25µL each) were separated by 12% SDS-PAGE, transferred to PVDF membrane, and detected via anti-c-Myc antibody (Part No. MA1-980), goat anti-mouse secondary antibody, and chemiluminescent substrate (Part No. 34080). Note: Manufacturers supply magnetic beads slurries at different concentrations and recommend different amounts of beads per microgram of lysate for IP. For this reason, only 10µL of CST beads were used in this experiment. Nevertheless, Pierce beads provide equal or greater binding capacity, even in experiments involving carefully matched numbers of beads (data not shown). Co-IP of BCL2-Luc with c-Myc-tagged BAD in 293T cellsThis experiment tests the specificity of co-immunoprecipitation by comparing results from 293T cells cultured alone (293T) or with vectors expressing BCL2-Luc and/or BAD-c-Myc fusion proteins (labeled BCL2 and BAD, respectively). The Western blot reveals the presence of BCL2 detected through its fused luciferase domain (i.e., via an anti-Luc antibody). Although BCL2 is expressed in two cultures (as indicated by its detection in two of the Control Lysates), it is purified by anti-c-Myc magnetic beads (IP or Co-IP lanes) only when co-expressed and captured through its interaction with c-Myc-tagged BAD (BAD+BCL2 lane). For the IP experiments, lysates were incubated with anti-c-Myc magnetic beads (50µL) for two hours at 4°C, then processed and eluted using the kit components and procedure. |
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