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His-tag

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    ​His-tags, or polyhistidine tags, comprise a consecutive series of six to ten histidine residues. The most common his-tag is the 6xHis tag and has a ​ molecular weight of 0.8kDa1. His tags are used for many recombinant proteins to facilitate purification, allowing researchers to extract a protein of interest from thousands of other proteins found in a cell or cell lysate. The small size of the 6x-His tag means there is a lower possibility for the tag to affect the functionality of the fusion protein, which can happen with larger tags. ​​

    Histidine forms coordination bonds with immobilized transition metal ions, and this property can be utilized for protein purification. His-tag protein purification is by a specialized form of affinity chromatography, called immobilized metal affinity chromatography (IMAC), where proteins or peptides are separated according to their affinity for metal ions immobilized to a solid chelating resin. During this process, a small His-tag is fused to either the N or C terminus of the target protein, enabling capture by nickel or cobalt ions coordinated on a variety of resins. The small size, low cost, and ease of use have made the His-tag the most popular affinity-tag available2. However, as His residues exist in mammalian and insect systems, there can be background binding. This can be avoided by washing in 5 – 10 mM imidazole, but this washing step may prematurely elute the protein of interest. 

    For more information on affinity tags, including the His-tag, see our guide for fusion tags.

    ​​

    Applications


    ​​Our range of antibodies, proteins, and assay kits are validated for use in various applications, including ELISA, western blot, immunohistochemistry, immunoprecipitation, immunocytochemistry, chromatin immunoprecipitation (ChIP), and flow cytometry.






    ​

    Immunoprecipitation of His/FLAG tagged PDCD5 using ab18184 at 1/100 dilution.

    Lane 1: MW Ladder (from top 120, 85, 50, 34, 25, 19 kDa)

    Lanes 2-5: Immunoprecipitation of His/FLAG tagged PDCD5 using ab18184 from mixture of ~50 ng purified His/FLAG tagged PDCD5 and 15 µL RRL in 300 µL buffer.

    Lanes 6-7: Purified PDCD5 from E. coli with ~60-70 ng protein.

    The lowest band in each lane is ~ 20 kDa and is PDCD5. The upper bands in lanes 2-5 are the heavy chain (~50 kDa, runs as doublet) and light chain (~25 or 26 kDa).

    Lane 1 : MW Ladder (from top 120, 85, 50, 34, 25, 19 kDa)
    Lanes 2-5 : Immunoprecipitation of His/FLAG tagged PDCD5 using ab18184 from mixture of ~50 ng purified His/FLAG tagged PDCD5 and 15 µL RRL in 300 µL buffer.
    Lanes 6-7 : Purified PDCD5 from E. coli with ~60-70 ng protein.

    Immunoprecipitation - Anti-6X His tag® antibody [HIS.H8] (ab18184)

    This image is courtesy of an Abreview submitted by Mr Chris Tracy




    Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HEK-293T (human epithelial cell line from embryonic kidney transformed with large T antigen) cells transfected with GFP-Myc-His vector expression construct labeling 6X His tag® with ab213204 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (white). Confocal image showing positive staining on HEK-293T cells transfected with GFP-Myc-His vector expression construct. The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) (ab195889) (red) at 1/200 dilution.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution.

    Immunocytochemistry/ Immunofluorescence - Anti-6X His tag® antibody [EPR20547] - ChIP Grade (ab213204)


    Product recommendations for your research




    Anti-6X His tag® antibodiesApplications
    Anti-6X His tag® antibody [HIS.H8] (ab18184)ICC/IF, ELISA, Dot blot, WB, IP, EMSA
    Anti-6X His tag® antibody (HRP) (ab1187)ELISA, WB
    Anti-6X His tag® antibody (ab9108)IP, WB, IHC-P, ICC/IF, ELISA, ChIP, Flow Cyt
    Anti-6X His tag® antibody (FITC) (ab1206)ELISA, WB, Flow Cyt
    Anti-6X His tag® antibody [EPR20547] (ab213204)WB, ICC/IF, IP, Flow Cyt, ChIP, IHC-P
    ELISA and Assay kitsSensitivity RangeDetection 
    GST 6XHis-tag ELISA Kit (ab128573)> 0.5 ng/mlColorimetric
    Protein

    6X His tag® peptide (ab14943)

    Protein Purification

    His-Tag Protein Purification Column (Pre-packed, 5 x 1ml) - AminTrap (ab270528)



    His-Tag Protein Purification Column (Pre-packed, 5 x 5ml) - AminTrap (ab270529)



    See our whole range of products to His-tag.


    References


    1. Malhotra, A. Tagging for Protein Expression Methods in Enzymology 463, Pages 239-258, (2009)
    2. Andersen, K. R., Leska, N. C., and Schwartz, T. U Optimized E. coli expression strain LOBSTR eliminates common contaminants from His‐tag purification PROTEINS: Structure, Functions, and Bioinformatics 81, Pages 1857-1861 (2013)






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