Epitope Tag Antibody Sampler Kit

• 产品编号：CST-9962S      品牌：Cell Signaling Technology       原厂货号：9962S
• 产品分类：抗体 > 一抗 > 复合抗体试剂盒
• 应用分类：

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描述：

表位标记抗体样本试剂盒中的所有抗体可以用于检测包含有相应表位标签的过表达的融合蛋白。DYKDDDK标签抗体识别DYKDDDK肽段（与Sigma公司Anti-FLAG®抗体识别的表位一致）并且它结合的特异性不依赖于存在的二价金属阳离子。鼠源的单克隆抗体是通过用6×His合成肽段（KLH-偶联）或者包含有流感病毒血凝素表位（YPYDVPDYA）（KLH-偶联）的合成肽段免疫小鼠生产的。兔源的单克隆抗体是通过用与人c-Myc（EQKLISEEDL）410-419残基相对应的合成肽段（KLH-偶联）或者GST融合蛋白免疫兔生产的。多克隆抗体是通过用合成DYKDDDK肽段（KLH-偶联）免疫兔生产的。多克隆抗体是采用蛋白A和肽亲和色谱方法提纯的。表位标记抗体样本试剂盒为分析多种多样的表位标记蛋白的表达提供了一种经济的研究方法。本试剂盒包括足量的一抗和二抗来做四次western miniblots实验 /一抗.在免疫印迹、免疫沉淀和免疫染色等技术中，表位标签用于标记和检测蛋白。由于它们的体积很小，它们一般不会影响标记蛋白的生化特性。目前，几种不同的表位标签已经得到广泛的应用并且易于制备。例如，很多质粒包含有编码一个氨基酸末端标签的DNA，该标签包括六个组氨酸（6×His）残基，其后为一个扩展的多克隆位点。已表达的重组蛋白上的6×His标签可以实现与亲和树脂的有效偶联和采用单步色谱纯化（1）。如同其它蛋白标签的系统(2),这种多组氨酸标签通常可以被蛋白酶如凝血酶和肠激酶在识别位点剪切,将目的蛋白分离(1)。谷胱甘肽巯基转移酶（GST）是另一种应用广泛的融合蛋白，因为它提供了一个易于检测的标签和简单的纯化过程，而又不影响目的蛋白的生物功能。在过去的十年里，大量带有GST-Tag的载体在原核和真核系统中被广泛开发和应用（3-5）。HA标签来源于流感病毒血凝素蛋白的一个表位，已经被作为一个常规表位标签，广泛应用于表达载体中（6），而Myc表位标签通常用来检测细菌、酵母、昆虫和哺乳动物细胞系统中融合蛋白的表达（7）。最后，DYKDDDK肽段也作为一种常规表位标签被广泛应用在表达载体中，并且它只包含8个氨基酸。这种肽段可以通过与氨基末端或羧基末端融合与目的蛋白一起被表达和检测到（8）。

1. Kroll, D.J. et al. (1993) DNA Cell Biol. 12, 441-453.
2. di Guan, C. et al. (1988) Gene 67, 21-30.
3. Guan, K.L. and Dixon, J.E. (1991) Anal. Biochem. 192, 262-267.
4. Davis, A.H. et al. (1993) Biotechnology 11, 933-936.
5. Yu, J. et al. (1998) Mol. Cell. Biol. 18, 1379-1387.
6. Field, J. et al. (1988) Mol. Cell. Biol. 8, 2159-2165.
7. Munro, S. and Pelham, H.R. (1984) EMBO J. 3, 3087-3093.
8. Brizzard, B. L. et al. (1994) Biotechniques 16, 730-735.

原厂资料：

Specificity / Sensitivity

All antibodies in the Epitope Tag Antibody Sampler Kit detect overexpressed fusion proteins containing the corresponding epitope tags. DYKDDDDK Tag Antibody recognizes the DYKDDDDK peptide (the same epitope recognized by Sigma's Anti-FLAG® antibodies), and its binding specificity is NOT dependent on the presence of divalent metal cations.

Source / Purification

Mouse monoclonal antibodies are produced by immunizing mice with a 6xHis synthetic peptide or a synthetic peptide containing the influenza hemagglutinin epitope (YPYDVPDYA) . Rabbit monoclonal antibodies are producted by immunizing rabbits with a synthetic peptide corresponding to residues 410-419 of human c-Myc (EQKLISEEDL) or with a GST fusion protein. Polyclonal antibodies are produced by immunizing animals with a synthetic DYKDDDDK peptide. Polyclonal antibodies are purified by protein A and peptide affinity chromatography.

Description

The Epitope Tag Antibody Sampler Kit provides an economical means to analyze the expression of a variety of epitope tagged proteins. The kit contains enough primary and secondary antibodies to perform four Western blots per primary antibody.

Background

Epitope tags are useful for the labeling and detection of proteins using immunoblotting, immunoprecipitation, and immunostaining techniques. Due to their small size, they are unlikely to affect the tagged protein's biochemical properties.Several different epitope tags are now commonly utilized and readily available. For instance, a variety of plasmids contain DNA that encodes an amino-terminal tag consisting of six histidine (6xHis) residues followed by an extended multiple cloning site. The 6xHis tag on the expressed recombinant proteins allows for efficient coupling to Ni2+ affinity resins and purification by single step chromatography (1). As is the case with other protein tag systems (2), this polyhistidine tag can often be cleaved at sites recognized by proteases such as thrombin and enterokinases to isolate the protein of interest (1). Glutathione S-transferase (GST) is another widely used fusion partner, since it provides both an easily detectable Tag and a simple purification process with little effect on the biological function of the protein of interest. Numerous vectors containing GST-Tag have been developed for both prokaryotic and eukaryotic systems over the past decade (3-5). The HA tag, derived from an epitope of the influenza hemagglutinin protein, has also been extensively used as a general epitope tag in expression vectors (6), while the Myc epitope tag is routinely used to detect expression of recombinant proteins in bacteria, yeast, insect and mammalian cell systems (7). Finally, the DYKDDDDK peptide has been used extensively as a general epitope tag in expression vectors and consists of only eight amino acids. This peptide can be expressed and detected with the protein of interested as an amino-terminal or carboxy-terminal fusion (8).

1. Kroll, D.J. et al. (1993) DNA Cell Biol. 12, 441-453.
2. di Guan, C. et al. (1988) Gene 67, 21-30.
3. Guan, K.L. and Dixon, J.E. (1991) Anal. Biochem. 192, 262-267.
4. Davis, A.H. et al. (1993) Biotechnology 11, 933-936.
5. Yu, J. et al. (1998) Mol. Cell. Biol. 18, 1379-1387.
6. Field, J. et al. (1988) Mol. Cell. Biol. 8, 2159-2165.
7. Munro, S. and Pelham, H.R. (1984) EMBO J. 3, 3087-3093.
8. Brizzard, B. L. et al. (1994) Biotechniques 16, 730-735.

注意事项：

Storage: Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM
NaCl, 100 µg/ml BSA and 50% glycerol. Store at –20°C. Do not
aliquot the antibodies.