Anti-Myc tag 抗体 [9E10] - ChIP Grade (ab32)

製品の概要

  • 製品名
    Anti-Myc tag antibody [9E10] - ChIP Grade
    Myc tag 一次抗体 製品一覧
  • 製品の詳細
    Mouse monoclonal [9E10] to Myc tag - ChIP Grade
  • 特異性
    This antibody is specific for Myc tagged proteins. The Myc tag epitope (EQKLISEEDL) is located at the dimerization site of c-myc and therefore this antibody does not perform well at recognizing endogenous c-myc. A publication by Baker AM et al. 2016 (PMID: 26826706 DOI: 10.1111/his.12939) shows the IHC staining generated by the 9E10 clone does not correlate with c-myc mRNA expression.
  • アプリケーション
    適用あり: ICC/IF, ICC, ChIP, IHC (Methanol fixed), Flow Cyt, WB, IP, ELISA, IHC-P, IHC-Fr, Purificationmore details
  • 免疫原

    Synthetic peptide corresponding to Human Myc tag aa 400-500 (C terminal) conjugated to keyhole limpet haemocyanin.

  • エピトープ
    aa 410-419 of human Myc.
  • ポジティブ・コントロール
    • Myc tagged proteins and myc tag expressing cells.
  • 特記事項

    If you require this antibody in a particular buffer formulation or a particular conjugate for your experiments, please contact orders@abcam.com or you can find further information here.

製品の特性

アプリケーション

Our Abpromise guarantee covers the use of ab32 in the following tested applications.

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

アプリケーション Abreviews 特記事項
ICC/IF Use a concentration of 5 µg/ml.
ICC 1/2000.

See Abreviews.

ChIP Use at an assay dependent concentration.
IHC (Methanol fixed) 1/200. PubMed: 17329357
Flow Cyt Use 1µg for 106 cells.

ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.

WB 1/500 - 1/1000.
IP Use at 6 µg/mg of lysate.
ELISA Use at an assay dependent concentration.
IHC-P 1/250 - 1/500.
IHC-Fr 1/1000. See Abreviews.
Purification Use at an assay dependent concentration.

ターゲット情報

  • 関連性
    Epitope tags are short peptide sequences that are easily recognized by tag-specific antibodies. Due to their small size, epitope tags do not affect the tagged protein’s biochemical properties. Most often sequences encoding the epitope tag are included with target DNA at the time of cloning to produce fusion proteins containing the epitope tag sequence. This allows anti-epitope tag antibodies to serve as universal detection reagents for any tag containing protein produced by recombinant means. This means that anti-epitope tag antibodies are a useful alternative to generating specific antibodies to identify, immunoprecipitate or immunoaffinity purify a recombinant protein. The anti-epitope tag antibody is usually functional in a variety of antibody-dependent experimental procedures. Expression vectors producing epitope tag fusion proteins are available for a variety of host expression systems including bacteria, yeast, insect and mammalian cells.
  • 細胞内局在
    Nuclear
  • 別名
    • c-myc tag antibody
    • Myc Epitope Tag antibody

画像

  • (A-D) Representative examples of body forming AtMORC7-MYC, AtMORC4-MYC, At-MORC1-MYC, and AtMORC6-MYC nuclei, respectively. (E) Untransformed wt nucleus subjected to the same antibody staining and imaging procedure. Left panels = anti-MYC channel; middle panels = DAPI channel (gray scaled). DAPI stains DNA, defining the position of dense chromocenters as high intensity white foci; right panels = merged channels (DAPI in blue, MYC in green). White triangles indicate examples of chromocenter adjacent AtMORC localization. Scale bars = 5 μM.

    Leaves from three-week old plants were fixed in 4% paraformaldehyde in TRIS buffer (10 mM TRIS pH 7.5, 10 mM EDTA, and 100 mM NaCl) for 20 minutes and washed twice in TRIS buffer. Leaves were chopped in 200–400 microliters lysis buffer (15 mM TRIS pH 7.5, 2 mM EDTA, 0.5 mM spermine, 80 mM KCl, 20 mM NaCl, and 0.1% Triton X-100) and filtered through a 3 μM cell strainer. 5 μL of nuclei suspension was added to 12 μL of sorting buffer (100mM TRIS pH 7.5, 50mM KCl, 2mM MgCl2, 0.05% Tween-20, and 20.5% sucrose) and air dried on chloroform dipped microscope slides for two hours and then post-fixed in 4% paraformaldehyde in PBS for 20 minutes. Slides were washed three times in PBS and incubated in blocking buffer (3% BSA, and 10% horse serum in PBS) for 30 minutes at 37°C. Nuclei were incubated at 4°C overnight in mouse monoclonal antibody against c-Myc (9E10, ab32; 1/200). Slides were washed in PBS and incubated with goat anti-mouse FITC antibody (ab7064; 1/200) for 90 minutes at room temperature. Following PBS washes, nuclei were counterstained and mounted in Vectashield mounting media with DAPI. Nuclei were analyzed with a Zeiss LSM 710 Confocal microscope at 63X or 100X magnification using Zen software.

     

  • Anti-Myc tag antibody [9E10] - ChIP Grade (ab32) at 1 µg/ml + E. coli Positive Control (Escherichia coli ) Whole Cell Lysate (ab5395) at 10 µg

    Secondary
    Goat polyclonal to Mouse IgG - H&L - Pre-Adsorbed (HRP) at 1/3000 dilution
    Developed using the ECL technique

    Performed under reducing conditions.

    Predicted band size : 41 kDa
    Observed band size : 45 kDa (why is the actual band size different from the predicted?)


    Exposure time : 1 minute

    Lysate from E. coli recombinantly expressing 11 commonly used tags including myc tag.

  • Ab32 staining a Myc tagged protein in HeLa cells by ICC/IF (Immunocytochemistry/Immunofluorescence). Endogenous c-myc was not detected under these conditions. Cells were fixed with paraformaldehyde, permeabilized with 0.5% Triton and blocked with 5% Serum for 30 minutes at 25°C. Samples were incubated with primary antibody (1/1000 in 5% Serum) for 1 hour at 25°C. An Alexa Fluor® 488 conjugated Goat anti-Mouse was used as a secondary antibody.

    See Abreview

  • RPE1 cells grown on coverslips were transfected with βarr2-myc, grown in low serum and then fixed and stained for Kif3A (red) and ab32 (green). Insets show higher magnifications of a representative PC. Kif3A was found in the cytoplasm and at the tip of the axoneme where it was colocalized with βarr2.

    Cells were incubated with primary antibodies in permeabilization buffer (PBS with 1 mg/mL bovine serum albumin (PBS-BSA) and 0.1% triton-X-100) for 45 minutes at room temperature. After two washes with PBS-BSA, cells were incubated for 30 minutes at room temperature in PBS-BSA containing secondary antibodies. After one wash with PBS-BSA and two washes in PBS, cells were laid down on microscope slides in a PBS–glycerol mix (50/50) with DAPI.



  • Predicted band size : 41 kDa

    Menssen R et al., (2001) Curr Biol. Mar 6;11(5):345-50.

    Phosphorylation of Cdc15 changes during the cell cycle. Exponentially growing cells (cyc) of CDC15-MYC9 (W1114) were arrested in G1 with a factor pheromone and released into fresh medium at 25°C. Cells were harvested at the indicated times, the percentage of divided nuclei was determined by DAPI staining of fixed cells, and proteins were analyzed by western blotting with 9E10 (ab32).

  • Cse4 histone variant occupancy at the endogenous CEN3 locus (eCEN3) and at a conditional CEN3 locus (cCEN3) when it is repressed (in the presence of galactose) or induced (glucose) by ChIP. Control (SC138) and Δfun30 (SC140) cells are grown in YP Gal until≈1 OD, then shifted in YP Glu or YP Gal containing 15 µg/ml of nocodazole and incubated for 4 hours. Cse4-Myc associated chromatin was immunoprecipitated and the immunoprecipitated DNA was analyzed by PCR followed by agarose gel electrophorsis and ethidium bromide staining to visualize the DNA fragments. Shown are the relevant, cropped out bands from a single, representative gel, input was 1/243th of the immunoprecipitated material.

    Overnight cultures grown in YPD at 30°C were diluted to 0.2 OD595, then grown to 0.7 OD595 at 30°C before crosslinking. Samples were crosslinked 15 min for H3, 3Myc-Htz1 and Cse4-Myc or 30 min for Fun30 detection with 1% final formaldehyde and chromatin extracts were sonicated to ∼500 bp. Triplicates or duplicate ChIP samples were validated by qPCR. Chromatin extracts were then immunoprecitated with 2 µg of mouse monoclonal anti-myc (9E10, ab32) for Cse4-Myc.

参考文献

This product has been referenced in:
  • Jiang H  et al. The regulator of calcineurin 1 increases adenine nucleotide translocator 1 and leads to mitochondrial dysfunctions. J Neurochem 140:307-319 (2017). WB ; Human . Read more (PubMed: 27861892) »
  • Cheung RS  et al. Ubiquitination-Linked Phosphorylation of the FANCI S/TQ Cluster Contributes to Activation of the Fanconi Anemia I/D2 Complex. Cell Rep 19:2432-2440 (2017). WB ; Human . Read more (PubMed: 28636932) »

See all 133 Publications for this product

レビューと Q&A

Application
ChIP
Sample
Human Cell lysate - whole cell (HEK cells)
Specification
HEK cells
Detection step
Real-time PCR
Type
Cross-linking (X-ChIP)
Duration of cross-linking step: 10 minute(s) and 0 second(s)
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投稿 Dec 12 2017

Application
CyTOF
Sample
Mouse Cell (Lung)
Specification
Lung
Username

Irit Milman

Verified customer

投稿 Nov 14 2017

Application
Western blot
Sample
Human Cell lysate - whole cell (HEK cells)
Gel Running Conditions
Reduced Denaturing (10)
Loading amount
30 µg
Specification
HEK cells
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: RT°C
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投稿 Nov 13 2017

Application
Immunocytochemistry/ Immunofluorescence
Sample
Human Cell (HeLa)
Permeabilization
Yes - 0.5% Triton
Specification
HeLa
Blocking step
Serum as blocking agent for 30 minute(s) · Concentration: 5% · Temperature: 25°C
Fixative
Paraformaldehyde
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投稿 Sep 05 2017

Application
Immunocytochemistry/ Immunofluorescence
Sample
Monkey Cell (Kidney)
Permeabilization
Yes - 0.1% Tween-20
Specification
Kidney
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 23°C
Fixative
Paraformaldehyde
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Dr. T Kidd

Verified customer

投稿 Jul 14 2017

Application
Western blot
Sample
Human Cell lysate - whole cell (HEK293)
Gel Running Conditions
Reduced Denaturing
Loading amount
10 µg
Specification
HEK293
Blocking step
Milk as blocking agent for 30 minute(s) · Concentration: 5% · Temperature: 25°C
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投稿 Jul 06 2016

Application
Western blot
Sample
Human Cell lysate - whole cell (LNCaP)
Gel Running Conditions
Reduced Denaturing (12)
Loading amount
40 µg
Specification
LNCaP
Blocking step
Odyssey Blocking Buffer as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 100% · Temperature: 26°C
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投稿 Jan 09 2015

Application
Western blot
Sample
Mouse Cell lysate - whole cell (Primary lung cancer line LKR10)
Gel Running Conditions
Reduced Denaturing
Loading amount
40 µg
Specification
Primary lung cancer line LKR10
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 2% · Temperature: 25°C
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投稿 Jul 25 2014

Application
Western blot
Sample
Human Cell lysate - whole cell (Lung cancer line A549)
Gel Running Conditions
Reduced Denaturing
Loading amount
40 µg
Specification
Lung cancer line A549
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 2% · Temperature: 25°C
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投稿 Jul 25 2014

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunoprecipitation
Sample
Human Cell lysate - whole cell (HeLa)
Total protein in input
20 µg
Immuno-precipitation step
Other - Magnetic Protein G
Specification
HeLa
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投稿 Jul 15 2013

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Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"

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