6X His Tag antibody

• Catalog Number: orb344413
• Category: Antibodies
• Description: 6X His Tag antibody
• Species/Host: Mouse
• Clonality: Monoclonal
• Clone Number: 33D10.D2.G8
• Tested applications: ELISA, FC, IHC, IP, WB
• Isotype: IgG1
• Immunogen: This His antibody was produced in mice by repeated immunizations with 6X His epitope tag peptide H-H-H-H-H-H conjugated to KLH using maleimide.
• Concentration: 1.0 mg/mL
• Dilution range: ELISA: 1:20,000 - 1:100,000, FC: User Optimized, IHC: User Optimized, IP: 1:200, WB: 1:2,000 - 1:10,000
• Form/Appearance: Liquid (sterile filtered)
• Purity: 6X His antibody was purified from concentrated tissue culture supernate by Protein A chromatography and is directed against the 6X His motif and is useful in determining its presence in various assays. This polyclonal anti-6X His tag antibody detects over-expressed proteins containing the 6X His epitope tag. To date, this antibody has reacted with all His tagged proteins so far tested. In western blotting of bacterial extracts, the antibody does not cross-react with endogenous proteins. The antibody recognizes the His-tag (His-His-His-His-His-His) fused to either the amino- or carboxy- termini of targeted proteins in transfected or transformed cells.
• Conjugation: Unconjugated
• Storage: Store 6X His Tag Antibody at -20° C or below prior to opening. This vial contains a relatively low volume of reagent (25 µL). To minimize loss of volume dilute 1:10 by adding 225 µL of the buffer stated above directly to the vial. Recap, mix thoroughly and briefly centrifuge to collect the volume at the bottom of the vial. Use this intermediate dilution when calculating final dilutions as recommended below. Store the vial at -20°C or below after dilution. Avoid cycles of freezing and thawing.
• Buffer/Preservatives: 0.01% (w/v) Sodium Azide
• Alternative names: mouse anti-6X His Tag Antibody, anti-HIS, HIS Anti
• Note: For research use only
• Application notes: Anti-6X His is optimally suited for monitoring expression of His-tagged fusion proteins. As such, Anti-6X His/6XHIS can be used to identify fusion proteins that contain the 6X His epitope. The antibody recognizes the His tag fused either to the amino- or carboxy- termini of targeted proteins. This antibody has been tested by ELISA, SDS-PAGE, and western blotting against both the immunizing peptide and His-containing recombinant proteins. Although not tested, this antibody is likely functional for immunoprecipitation and immunocytochemistry.
• Expiration Date: 12 months from date of receipt.

Characterization of purity and antigenicity of CKAP4 constructs. a SDS-PAGE of normalized amounts of purified CKAP4 constructs followed by silver staining. b–f Western blots using anti-His tag, anti-CKAP4 (G1/296), anti-CKAP4 (epitope region: 216–265), anti-CKAP4 (epitope region: 375–425), and anti-CKAP4 (epitope region: 552–602) primary antibody, respectively. Separate blots were used for each antibody.

ELISA of Mouse anti-6xHIS Tag Antibody. Antigen: HIS-tagged purified protein and E. coli cell lysates expressing HIS-Tagged construct, GST- and RON-tagged purified proteins. Coating amount: 0.15 ug per well. Primary antibody: 6xHIS Tag antibody at 100 ug/ml. Dilution series: 2-fold. Mid-point concentration: 200 ng/mL. Secondary antibody: Peroxidase mouse secondary antibody at 1:10000. Substrate: TMB (p/n orb348651).

Western Blot of Mouse Anti-6X HIS Epitope Tag Antibody. Lane 1: Opal Prestained Molecular Weight Marker. Lane 2: rec EGF -C-term HIS tag [0.02 µg] [+]. Lane 3: rec Luciferase N-term HIS tag [0.10 µg] [+]. Lane 4: Zic1 - C-myc/DDK Tag [0.10 µg] [-]. Primary Antibody: Mouse Anti-6X HIS at 1.0 µg/mL overnight at 2-8°C. Secondary Antibody: Rabbit Anti-Mouse HRP (p/n orb347506) at 1:40000 for 30 mins at RT. Block: BlockOut Buffer (p/m orb348644) for 1 hr at RT. Bands are recognized at the expected MW. Exposure: 2sec.

Western Blot of Mouse anti-6xHIS Tag Antibody. Lane 1: 100 ng Purified histidine-tagged recombinant protein. Lane 2: 200 ng E. coli cell lysate containing histidine-tagged expression construct. Lane 3: 100 ng Purified GST-tagged recombinant protein. Lane 4: 100 ng Purified FLAG-tagged recombinant protein. Primary antibody: Mouse anti-6xHIS Tag antibody at 1:5000 overnight at 4°C. Secondary antibody: Peroxidase mouse secondary antibody at 1:20000 for 30 min at RT. Block: 5% BLOTTO for 1 hr at RT.