Anti-6X His tag® antibody - ChIP Grade (ab9108)

Overview

  • Product nameAnti-6X His tag® antibody - ChIP Grade
    See all 6X His tag® primary antibodies
  • Description
    Rabbit polyclonal to 6X His tag® - ChIP Grade
  • SpecificityELISA: Antibody specificity was verified by ELISA against peptide conjugated to BSA (HHHHHH-BSA). A 1:25,000 dilution of the antibody gave an OD=1.0 in a 15 minute reaction using HRP-conjugated Goat anti-Rabbit IgG at 1:20,000 and TMB as the substrate. Appropriate specificity controls were run.
  • Tested applicationsSuitable for: IP, WB, IHC-P, ICC/IF, ELISA, ChIP, Flow Cytmore details
  • Immunogen

    Synthetic peptide:

    HHHHHH

    conjugated to KLH.

  • Positive control
  • General notesThis antibody recognizes both C-terminal and N-terminal tags. HIS-TAG® is a trademark of EMD Biosciences, Inc.

Properties

Applications

Our Abpromise guarantee covers the use of ab9108 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
IP Use at an assay dependent concentration.
WB 1/1000.
IHC-P Use at an assay dependent concentration. PubMed: 21483675
ICC/IF 1/100 - 1/500.
ELISA 1/1000.
ChIP Use at an assay dependent concentration. ChIP was performed with 25 ug chromatin, 5ug of antibody and 20 ul of Protein A/G beads.
Flow Cyt Use at an assay dependent concentration. (see Abreview published on 22nd January, 2007).



ab171870-Rabbit polyclonal IgG, is suitable for use as an isotype control with this antibody.

Target

  • RelevanceThe H-H-H-H-H-H motif is used as a tag on many recombinant proteins to facilitate purification. The antibody recognizes the His-tag fused to the amino- or carboxy- termini of targeted proteins in transfected or transformed cells.
  • Cellular localizationDepends upon the localization of the parent protein tagged with hexahistidine.
  • Alternative names
    • 6 His epitope tag antibody
    • Hexa His tag antibody
    • HHHHHH epitope tag antibody
    • HHHHHH tag antibody
    • His tag antibody
    • Polyhistidine Tag antibody
    see all

Anti-6X His tag® antibody - ChIP Grade images

  • A stably transfected 293T human cell line harbouring the GAL4 upstream activation sequence was transiently transfected with a Myc or His - tagged GAL4 DNA Binding Domain construct. 48 hours post transfection Chromatin was prepared according to the Abcam X-ChIP protocol. The ChIP was performed with 25 ug chromatin, 5ug of antibody and 20 ul of Protein A/G beads. A non-specific antibody was used as the negative control. The immunoprecipitated DNA was quantified by real time PCR (SYBR Green approach).
  • All lanes : Anti-6X His tag® antibody - ChIP Grade (ab9108) at 1/4000 dilution

    Lane 1 : HeLa whole cell lysate - transfected with empty vector
    Lane 2 : HeLa whole cell lysate - transfected with His-tagged gene

    Lysates/proteins at 10 µg per lane.

    Secondary
    HRP-conjugated goat anti-rabbit IgG polyclonal at 1/5000 dilution
    Developed using the ECL technique

    Performed under reducing conditions.

    Observed band size : 35 kDa (why is the actual band size different from the predicted?)


    Exposure time : 30 seconds

    This image is courtesy of an anonymous Abreview

    See Abreview

  • ab9108 staining CACNB3 with His tag expressed in HEK293 cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with paraformaldehyde, permeabilized with 0.5% Triton X-100 and blocked with 5% serum for 20 minutes at 25°C. Samples were incubated with primary antibody (1/500 in 1% goat serum, 0.1% TX100 and PBS) for 16 hours at 4°C. An  Alexa Fluor® 488-conjugated Goat anti-rabbit IgG polyclonal was used as the secondary antibody (1/750).

    See Abreview

  • Use of ab9108 in western blotting at 1:1000 against Epitope Tag Positive Control lysate (ab4524).
    Lane 1: 2 µl lysate
    Lane 2: 1 µl lysate
    Lane 3: 0.5 µl lysate
    Lane 4: 0.25 µl lysate Use of ab9108 in western blotting at 1:1000 against E.coli Positive Control (Escherichia coli ) Whole Cell Lysate - expressing 11 different epitope tag (ab5395). Lane 1: 2 µl lysate Lane 2: 1 µl lysate Lane 3: 0.5 µl lysate Lane 4: 0.25 µl lysate
  • All lanes : Anti-6X His tag® antibody - ChIP Grade (ab9108) at 1/1000 dilution

    Lane 1 : 10ug non-transformed lysate
    Lane 2 : 10ug non-transformed lysate + 1mM IPTG
    Lane 3 : 10ug E.coli Lysate harbouring His-tag-RelA NTD, non-induced
    Lane 4 : 10ug Induction of His-tag RelA NTD 1 mM IPTG
    Lane 5 : 10ug Induction of His-tag RelA NTD 1 mM IPTG

    Secondary
    HRP conjugated goat anti-rabbit IgG
    Developed using the ECL technique

    Performed under reducing conditions.

    Observed band size : 37 kDa (why is the actual band size different from the predicted?)
    Additional bands at : 12 kDa (possible non-specific binding).

    Exposure time : 1 minute
    This image is courtesy of an Abreview.

    See Abreview

References for Anti-6X His tag® antibody - ChIP Grade (ab9108)

This product has been referenced in:
  • Shigemori S  et al. Oral delivery of Lactococcus lactis that secretes bioactive heme oxygenase-1 alleviates development of acute colitis in mice. Microb Cell Fact 14:189 (2015). Mouse . Read more (PubMed: 26608030) »
  • Blanka A  et al. Identification of the alternative sigma factor SigX regulon and its implications for Pseudomonas aeruginosa pathogenicity. J Bacteriol 196:345-56 (2014). ChIP . Read more (PubMed: 24187091) »

See all 33 Publications for this product

Product Wall

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Sample African Green Monkey Recombinant protein (COS 7 cells)
Loading amount 10 µg
Specification COS 7 cells
Treatment Homogenised
Gel Running Conditions Reduced Denaturing (12% tris glycine)
Blocking step BSA as blocking agent for 16 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
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Submitted Mar 27 2013

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Sample Escherichia coli Cell lysate - whole cell (membrane pellet homogenised)
Loading amount 10 µg
Specification membrane pellet homogenised
Gel Running Conditions Reduced Denaturing (10-20% Tricine)
Blocking step BSA as blocking agent for 16 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
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Submitted Mar 12 2013

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunoprecipitation
Sample Human Cell lysate - other (HEK Cell memrane lysate incubated with supernatant)
Total protein in input 100000 cells
Specification HEK Cell memrane lysate incubated with supernatant
Immuno-precipitation step Protein G
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Submitted Feb 01 2013

Application ELISA
Sample Human Cell (NIH3T3)
Specification NIH3T3
Blocking step BSA as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 1% · Temperature: 25°C
Type Sandwich (Capture)
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Submitted Oct 21 2016

Application Immunoprecipitation
Immuno-precipitation step Protein G
Sample Human Cell lysate - whole cell (HeLa)
Specification HeLa
Treatment 100ug/ml FAC or 100uM DFO for 8hrs
Total protein in input 5 µg
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Submitted Apr 18 2014

Application Western blot
Loading amount 10 µg
Gel Running Conditions Reduced Denaturing (10)
Sample Human Cell lysate - whole cell (HeLa)
Specification HeLa
Treatment transfected with His-tagged gene
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 23°C
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Submitted Mar 29 2014

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Sample Mouse Recombinant protein (HEK Cell supernatant)
Loading amount 40 µg
Specification HEK Cell supernatant
Gel Running Conditions Non-reduced Denaturing (15)
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 2% · Temperature: 25°C
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Submitted Sep 13 2012

Unfortunately we do not have a His tagged BSA in our catalog and can not offer HIS-tagged BSA for sale as it's just a component within our QC procedures.

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Sample Human Cell lysate - whole cell (HEK293 cells)
Loading amount 20 µg
Specification HEK293 cells
Gel Running Conditions Non-reduced Denaturing (gel 12%)
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 24°C
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Abcam user community

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Submitted Nov 11 2011

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Sample Human Purified protein (His-tagged purified protein (histone H2B))
Loading amount 0.5 µg
Specification His-tagged purified protein (histone H2B)
Gel Running Conditions Reduced Denaturing (15%)
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 4% · Temperature: 25°C
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Dr. Efrat Shema

Verified customer

Submitted Aug 11 2011

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