Overview

  • Product name

  • Description

    Mouse polyclonal to ADAR1
  • Host species

    Mouse
  • Tested applications

    Suitable for: WB, IHC-P, ICC/IFmore details
  • Species reactivity

    Reacts with: Human
  • Immunogen

    Full length protein, corresponding to amino acids 1-1226 of Human ADAR1(P55265)

  • Positive control

    • WB: lysate from liver or HepG2 cells. IHC: lung tissue. ICC/IF: HeLa cells.

Properties

Applications

Our Abpromise guarantee covers the use of ab88574 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
WB Use a concentration of 1 µg/ml. Predicted molecular weight: 104 kDa.
IHC-P Use a concentration of 3 µg/ml. Antigen retrieval is not essential but may optimise staining.
ICC/IF Use a concentration of 10 µg/ml.

Target

  • Function

    Converts multiple adenosines to inosines and creates I/U mismatched base pairs in double-helical RNA substrates without apparent sequence specificity. Has been found to modify more frequently adenosines in AU-rich regions, probably due to the relative ease of melting A/U base pairs as compared to G/C pairs. Functions to modify viral RNA genomes and may be responsible for hypermutation of certain negative-stranded viruses. Edits the messenger RNAs for glutamate receptor (GLUR) subunits by site-selective adenosine deamination. Produces low-level editing at the GLUR-B Q/R site, but edits efficiently at the R/G site and HOTSPOT1. Binds to short interfering RNAs (siRNA) without editing them and suppresses siRNA-mediated RNA interference. Binds to ILF3/NF90 and up-regulates ILF3-mediated gene expression.
  • Tissue specificity

    Ubiquitously expressed, highest levels were found in brain and lung.
  • Involvement in disease

    Defects in ADAR are a cause of dyschromatosis symmetrical hereditaria (DSH) [MIM:127400]; also known as reticulate acropigmentation of Dohi. DSH is a pigmentary genodermatosis of autosomal dominant inheritance characterized by a mixture of hyperpigmented and hypopigmented macules distributed on the dorsal parts of the hands and feet.
  • Sequence similarities

    Contains 1 A to I editase domain.
    Contains 2 DRADA repeats.
    Contains 3 DRBM (double-stranded RNA-binding) domains.
  • Post-translational
    modifications

    Sumoylation reduces RNA-editing activity.
  • Cellular localization

    Cytoplasm. Nucleus > nucleolus. Isoform 1 is found predominantly in cytoplasm but appears to shuttle between the cytoplasm and nucleus. Isoform 5 is found exclusively in the nucleolus.
  • Information by UniProt
  • Database links

  • Alternative names

    • 136 kDa double-stranded RNA-binding protein antibody
    • 136kDa double stranded RNA binding protein antibody
    • Adar 1 antibody
    • ADAR antibody
    • Adar1 antibody
    • Adenosine deaminase acting on RNA 1 A antibody
    • Adenosine deaminase RNA specific 1 antibody
    • Adenosine deaminase RNA specific antibody
    • Adenosine deaminase that act on RNA antibody
    • AGS6 antibody
    • AV242451 antibody
    • Double stranded RNA specific adenosine deaminase antibody
    • Double-stranded RNA-specific adenosine deaminase antibody
    • Double-stranded RNA-specific editase Adar antibody
    • DRADA antibody
    • Dsh antibody
    • Dsrad antibody
    • DSRAD_HUMAN antibody
    • dsRNA adenosine deaminase antibody
    • EC 3.5.4.- antibody
    • G1P1 antibody
    • IFI 4 antibody
    • IFI-4 antibody
    • IFI4 antibody
    • Ifi4 protein antibody
    • Interferon induced protein 4 antibody
    • Interferon inducible protein 4 antibody
    • Interferon-inducible protein 4 antibody
    • K88DSRBP antibody
    • mZaADAR antibody
    • P136 antibody
    • Pre-mRNA adenosine deaminase antibody
    • RNA adenosine deaminase 1 antibody
    • RNA-editing deaminase 1 antibody
    • RNA-editing enzyme 1 antibody
    see all

Images

  • Anti-ADAR1 antibody (ab88574) at 1 µg/ml + Human liver lysate at 50 µg

    Predicted band size: 104 kDa

  • Immunofluorescence analysis of MOCK-infected HEK293T cells, staining ADAR1 (green) with ab88574.

    Cells were fixed in 4% formaldehyde and permeabilized with 0.2% Triton X-100 before incubating with fluorescent-conjugated anti-mouse secondary antibody. Nuclei were stained with DAPI (blue).
  • ab88574 at 3µg/ml staining ADAR1 in formalin-fixed, paraffin-embedded Human lung tissue.
  • Anti-ADAR1 antibody (ab88574) at 1 µg/ml + HepG2 cell lysate at 50 µg

    Predicted band size: 104 kDa

  • Immunofluorescent staining of ADAR1 on HeLa cells using ab88574 at 10 µg/ml.

References

This product has been referenced in:

  • Yu J  et al. ADAR1 p110 Enhances Adhesion of Tumor Cells to Extracellular Matrix in Hepatocellular Carcinoma via Up-Regulating ITGA2 Expression. Med Sci Monit 25:1469-1479 (2019). Read more (PubMed: 30798327) »
  • Shigeyasu K  et al. AZIN1 RNA editing confers cancer stemness and enhances oncogenic potential in colorectal cancer. JCI Insight 3:N/A (2018). Read more (PubMed: 29925690) »
See all 13 Publications for this product

Customer reviews and Q&As

1-5 of 5 Abreviews or Q&A

Application
Immunocytochemistry/ Immunofluorescence
Sample
Human Cell (HEK-293)
Permeabilization
Yes - 0.3% Triton X-100 in Blocking Buffer
Specification
HEK-293
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Fixative
Paraformaldehyde

Abcam user community

Verified customer

Submitted Mar 22 2018

Application
Immunoprecipitation
Sample
Human Cell lysate - whole cell (Breast Cancer Cell Lines (MCF7 & BT549))
Total protein in input
6e+007 cells
Immuno-precipitation step
Protein G
Specification
Breast Cancer Cell Lines (MCF7 & BT549)

Abcam user community

Verified customer

Submitted Apr 27 2016

Answer

The ADAR antibody ab88574 is raised against full-length human ADAR1, and it is a polyclonal antibody, which means it will potentially recognize all ADAR isoforms.

Polyclonal antibodies are composed of many antibodies specific for different epitopes (amino acid sequences) on the immunogen (full-length ADAR, in this case). Monoclonal antibodies are specific for a single epitope.

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Question
Answer

Thank you for your enquiry.

I am pleased to provide the following IHC-P protocol used for testing ab88574 . As discussed on the telephone, please note that this would be a guideline only and may require some further optimization.

I hope this will be helpful. If you have any further questions, please do not hesitate to contact us.



Immunohistochemistry Method

Deparaffinize sections and rehydrate using PBS.

Pre-treat (antigen retrieval) the sample with one of the following methods:

No treatment at all.
Place sample in 1X citrate buffer (pH 6.0) in pressure cooker under 125℃
for 4min and under 90℃ for 45min, cool sample subsequently.

Place sample in 1X citrate buffer (pH 6.0) and microwave at 750W for 20 minutes, cool
sample subsequently.
Place sample in 1X Tris/EDTA buffer (pH 9.0) and microwave at 750W for 20 minutes,
cool sample subsequently.
Place sample in HCl (2N) (pH 0.6˜0.9) at room temperature for 10˜20 minutes.
Place sample in 0.1% trypsin and shake for 25 minutes at 37°C.

Step-by-step procedure:
1. Incubate sections in 3% H2O2 in 1X PBS at room temperature for 10 minutes and then
wash the sections again.
2. Incubate sections in blocking solution for 10 minutes.
3. Add primary antibodies (diluted in blocking solution) and incubate the sections overnight at
4°C, wash sample with 1X PBS afterwards.
4. Incubate sections with labeled polymer for 30 min followed by washing the sections with
PBS.
5. Application of substrate solution (DAB or other suitable peroxidase substrate). Wash
sample thoroughly under running tap water.
6. Counter stain the samples in Mayer’s hematoxylin.
7. Dehydrate and mount samples.

Reagents:

Blocking solution or Antibody Diluent (commercially purchased)
Immunodection Kit- EnVision Detection Kit, Peroxidase/DAB,Rabbit/Mouse
Citrate buffer, pH 6.0: 10 mM sodium citrate buffer
1X Tris/EDTA, pH 9.0: 10 mM Tris base, 1 mM EDTA solution, 0.05% Tween 20
HCl solution (2N), pH 0.6˜0.9: prepared in distilled water

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Question

We recently ordered antibody ab97640 against ADAR1 from Abcam. We have tried a number of experiments to detect both endogenous and overexpressed human ADAR1 by western blot and immunofluorescence, and we find that the antibody does not work at all. Specifically, we performed a western blot of lysates from HEK293T cells in which we over expressed either isoform of human ADAR1; these isoforms were tagged with a c-terminal V5 tag. The ADAR1 antibody, used at the recommended concentration for western blot, did not detect any endogenous ADAR1 in the untransfected 293T cell lysates, nor did it detect either of the over expressed isoforms of human ADAR1. We stripped and reprobed this blot with V5 antibody and found abundant expression of our tagged ADAR1 constructs, so the proteins were indeed expressed properly in this experiment. In parallel, we attempted to detect endogenous and over-expressed ADAR1 by immunofluorescence on paraformaldehyde- fixed HeLa cells. Again, the V5 staining worked for the tagged, overexpressed proteins, but the Abcam antibody did not work at all for either the endogenous or the overexpressed proteins. I noticed that there was a recent Abreview in which another group found that the antibody did not work at all. The Abcam response was that "It seems that it is a vial problem. We have not had similar complaints." I wonder if our antibody came from the same lot as the one mentioned in the review. We would like to exchange this antibody for another Abcam antibody to ADAR1 that has a history of working properly: ab88574.

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Answer

Thank you for contacting Abcam about this issue. I am sorry this product did not perform as stated on the datasheet and for the inconvenience this has caused. Before I may issue a free of charge replacement, I would appreciate it if you could send me the specific of the protocols that you had used with Ab97640 for our records. The details provided enable us to closely monitor the quality of our products.   For your Western blotting protocol could you please let me know about your blocking step such as what you blocked in, for how long and at what temperature? Could you also inform us as to the incubation conditions for the primary and secondary antibodies? What concentrations these were used at, as well as any adustments you made while troubleshooting? Where you able to run an isotype control? For your ICC-IF could you give me the same information as well as antigen retrieval steps you may have used? Thank you very much for your cooperation. If you have any questions please feel free to contact us.

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