Western Blotting Using In-Gel Protein Labeling as a Normalization Control: Stain-Free Technology
Western Blotting Using In-Gel Protein Labeling as a Normalization Control: Stain-Free Technology
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DOI：
10.1007/978-1-4939-2550-6_27
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发表时间：
2015-01-01
期刊：
影响因子：
--
通讯作者：
Gomes, Aldrin V.
中科院分区：
文献类型：
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作者：
Gilda, Jennifer E.;Gomes, Aldrin V.
蛋白质印迹法是一种常用的实验室技术，用于半定量蛋白质量。当定量蛋白质表达时，重要的是使用上样对照来解释上样到凝胶上的总蛋白质的量的差异。常见的上样对照包括管家蛋白，如β-肌动蛋白或GAPDH，通过蛋白质印迹定量，或总蛋白，使用染色剂如考马斯亮蓝或丽春红S定量。最近开发的总蛋白质定量方法利用免染色技术，其具有线性动态检测范围，并允许在凝胶和膜上进行蛋白质检测。在这里，我们描述的理论和使用免染色凝胶的总蛋白定量和标准化的蛋白质印迹。
Western blotting is a commonly used laboratory technique for semi-quantifying protein amounts. It is important when quantifying protein expression to account for differences in the amount of total protein loaded onto the gel using a loading control. Common loading controls include housekeeping proteins, such as beta-actin or GAPDH, quantified by Western blot, or total protein, quantified using a stain such as Coomassie Brilliant Blue or Ponceau S. A more recently developed method for total protein quantification utilizes stain-free technology, which has a linear dynamic detection range and allows for protein detection on both gels and membranes. Here, we describe the theory and use of stain-free gels for total protein quantification and normalization of Western blots.