6x-His Tag Monoclonal Antibody (4E3D10H2/E3)/His标签抗体

货号：MA1-135

规格：100 ug

价格：2753

产品类型：标签抗体

品牌：Thermo Fisher

物种：Tag

宿主：小鼠

抗体亚型：其它

荧光染料：其它

点击Invitrogen标签抗体集，查看更多标签抗体

抗体类型:	单抗	同型对照：	IgG1
免疫原性：	Mixture of N-terminal and C-terminal 6x-His peptides, CG-HHHHHH and HHHHHH-GC	用法：	25-500 ng/mL (ELISA);2 µg（IP); 1:250-1:1000（WB)

Product Specific InformationBy Western blot, MA1-135 detects both N-terminal and C-terminal His-tagged proteins.Background/Target InformationThe 6x His tag is a synthetic oligo peptide consisting of 6 consecutive histidine residues (HHHHHH). The His tag is commonly expressed as a tag at either N-or C-terminal regions of recombinant proteins to allow isolation or purification by immobilized metal affinity chromatography. Epitope tagging is a technique in which a known epitope is fused to a recombinant protein using genetic engineering. By choosing a particular epitope and recombinant protein combination, epitope tagging makes it possible to detect proteins for which no antibody is available. His epitope tagged proteins that contain a stretch of histidine residues at the carboxyl or amino terminus are used for purification of recombinant proteins by means of metal chelate chromatography due to metal binding properties of histidine clusters on His tagged proteins. For example, the affinity of the histidine-tag motif to Ni2+ by chelation is strong and selective enough to enable purification of the protein to homogeneity by affinity chromatography on a Ni2+-NTA adsorbent. His tag-specific antibodies are used to facilitate detection or coimmunopreciptation of tagged proteins.For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.

数据

6x-His Tag Antibody (MA1-135) in ELISADirect ELISA analysis of N-terminal and C-terminal His-tagged proteins was performed by coating wells of a plate with a recombinant N-terminal His-tagged protein (left panel) or a recombinant C-terminal His-tagged protein (right panel) at a concentration of 5 µg/mL overnight at 4C. The plate was washed 3 times with ELISA Wash Buffer (Product # N503), 100 µL of a 6x-His Epitope Tag monoclonal antibody (Product # MA1-135) was added to wells in duplicate at 1250, 625, 312.5, 156, 78, 39, 19.5, 9.5, 4.5 and 0 ng/mL concentrations, and the samples were incubated for 2 hours at room temperature. The plate was washed, and then incubated with 100 µL per well of an HRP-conjugated goat anti-mouse IgG secondary antibody (Product # 31430) at a dilution of 1:10,000 for 1 hour at room temperature, and washed again with ELISA Wash Buffer. The plate was developed by incubating 100 µL per well of 1-Step Ultra TMB substrate (Product # 34028) per well for 10 minutes at room temperature in the dark. The reaction was stopped with 100 µL per well of 0.16M sulfuric acid. Absorbances were read on a spectrophotometer at 450-550 nm. 6x-His Tag Antibody (MA1-135) in IPImmunoprecipitation of recombinant His-tagged proteins was performed on E. coli cell lysates expressing N-terminal and C-terminal His-tagged GFP. Antigen-antibody complexes were formed by incubating 1 mg of whole cell lysate with 5 µg of a 6x-His Epitope Tag monoclonal antibody (Product # MA1-135) overnight on a rocking platform at 4C. The immune complexes were captured on 50 µL Protein A/G Agarose (Product # 20421), washed extensively, and eluted with Lane Marker Reducing Sample Buffer (Product # 39000). Samples were resolved on a 4-20% Tris-HCl polyacrylamide gel, transferred to a PVDF membrane, and blocked with StartingBlock (PBS) Blocking Buffer (Product # 37538) for at least 1 hour at room temperature. N-terminal His-tagged GFP was detected at ~37 kD and C-terminal His-tagged GFP was detected at ~28 kD using a 6x-His Epitope Tag monoclonal antibody (Product # MA1-135) at a concentration of 2 µg/mL in blocking buffer overnight at 4C on a rocking platform, followed by an HRP-conjugated goat anti-mouse IgG (Fc) secondary antibody (Product # 31439) at a dilution of 1:20,000 for at least 1 hour. Chemiluminescent detection was performed using SuperSignal West Pico (Product # 34080). 6x-His Tag Antibody (MA1-135) in WBWestern blot analysis of recombinant N-terminal His-tagged turbo GFP was performed by loading the indicated amounts of protein, and 15 µL of PageRuler Prestained Protein Ladder (Product # 26616) per well onto a 4-20% Tris-HCl polyacrylamide gel. Proteins were transferred to a PVDF membrane using the G2 Fast Blotter (Product # 62288) and blocked with StartingBlock (PBS) Blocking Buffer (Product # 37538) for at least 1 hour at room temperature. N-terminal His-tagged turbo GFP was detected at ~37 kD using a 6x-His Epitope Tag monoclonal antibody (Product # MA1-135) at a concentration of 2 µg/mL in blocking buffer overnight at 4C on a rocking platform, followed by an HRP-conjugated goat anti-mouse IgG (Fc) secondary antibody (Product # 31439) at a dilution of 1:20,000 for at least 1 hour. Chemiluminescent detection was performed using SuperSignal West Pico (Product # 34080). 6x-His Tag Antibody (MA1-135) in WBWestern blot analysis of recombinant C-terminal His-tagged GFP-2 was performed by loading the indicated amounts of protein, and 15 µL of PageRuler Prestained Protein Ladder (Product # 26616) per well onto a 4-20% Tris-HCl polyacrylamide gel. Proteins were transferred to a PVDF membrane using the G2 Fast Blotter (Product # 62288) and blocked with StartingBlock (PBS) Blocking Buffer (Product # 37538) for at least 1 hour at room temperature. C-terminal His-tagged GFP-2 was detected at ~28 kD using a 6x-His Epitope Tag monoclonal antibody (Product # MA1-135) at a concentration of 2 µg/mL in blocking buffer overnight at 4C on a rocking platform, followed by an HRP-conjugated goat anti-mouse IgG (Fc) secondary antibody (Product # 31439) at a dilution of 1:20,000 for at least 1 hour. Chemiluminescent detection was performed using SuperSignal West Pico (Product # 34080).

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参考文献：

1.Nature communications Design of Peptoid-peptide Macrocycles to Inhibit the β-catenin TCF Interaction in Prostate Cancer. View published figure on By clicking the link above, you will be redirected to the Benchsci website.Disclaimer

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