Overview

  • Product name
    Donkey Anti-Mouse IgG H&L (HRP)
    See all IgG secondary antibodies
  • Host species
    Donkey
  • Target species
    Mouse
  • Specificity
    The antibody used for conjugation reacts with mouse immunoglobulins of all classes. Cross-reactions as determined by ELISA for the unconjugated antibody (ab182022): Human IgG, rabbit IgG, goat IgG and Chicken IgY, less than 2%. Rat IgG, less than 33%.
  • Tested applications
    Suitable for: WB, IP, ELISA, IHC-Pmore details
  • Immunogen

    The details of the immunogen for this antibody are not available.

  • Conjugation
    HRP

Properties

  • Form
    Liquid
  • Storage instructions
    Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C. Avoid freeze / thaw cycle. Store In the Dark.
  • Storage buffer
    pH: 7.4
    Preservative: 0.1% Proclin
    Constituents: PBS, 1% BSA, 30% Glycerol
  • Concentration information loading...
  • Purity
    Immunogen affinity purified
  • Purification notes
    This antibody was isolated by affinity chromatography using antigen coupled to agarose beads and conjugated to Horse Radish Peroxidase (HRP).
  • Clonality
    Polyclonal
  • Isotype
    IgG
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab205724 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 1/2000 - 1/20000.
IP Use at an assay dependent concentration.
ELISA Use at an assay dependent concentration.
IHC-P 1/2000 - 1/20000.

Images

  • IHC image of Histone H4 staining in a section of formalin-fixed paraffin-embedded normal human colon*. The section was pre-treated using pressure cooker heat mediated antigen retrieval with sodium citrate buffer (pH6) for 30mins, and incubated overnight at +4°C with ab31830 at 0.5ug/ml. DAB was used as the chromogen (ab103723), diluted 1/100 and incubated for 10min at room temperature.

    An HRP-conjugated secondary (Ab205724, 1/2000 dilution) was used for 1hr at room temperature.

    The section was counterstained with haematoxylin and mounted with DPX. The inset negative control image is taken from an identical assay without primary antibody.

    For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

    *Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

  • All lanes : Anti-beta Actin antibody [mAbcam 8226] - Loading Control (ab8226) at 1 µg/ml

    Lane 1 : Liver (Human) Tissue Lysate
    Lane 2 : Liver (Mouse) Tissue Lysate
    Lane 3 : Liver (Rat) Tissue Lysate
    Lane 4 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate
    Lane 5 : NIH 3T3 (Mouse embryonic fibroblast cell line) Whole Cell Lysate
    Lane 6 : PC12 (Rat adrenal pheochromocytoma cell line) Whole Cell Lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : Donkey Anti-Mouse IgG H&L (HRP) (ab205724) at 1/2000 dilution

    Developed using the ECL technique.

    Performed under reducing conditions.

    Observed band size: 42 kDa
    why is the actual band size different from the predicted?


    Exposure time: 15 seconds


    This blot was produced using a 4-12% Bis-tris gel under the MOPS buffer system. The gel was run at 200V for 50 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour using 2% Bovine Serum Albumin before being incubated with ab8226 overnight at 4°C. Antibody binding was detected using ab205724, and visualised using ECL development solution ab133406.

  • IHC image of alpha tubulin staining in a section of formalin-fixed paraffin-embedded normal human colon*. The section was pre-treated using pressure cooker heat mediated antigen retrieval with sodium citrate buffer (pH6) for 30mins, and incubated overnight at +4°C with ab7291 at 0.5ug/ml.

    An HRP-conjugated secondary (Ab205725, 1/2000 dilution) was used for 1hr at room temperature. DAB was used as the chromogen (ab103723), diluted 1/100 and incubated for 10min at room temperature. The section was counterstained with haematoxylin and mounted with DPX. The inset negative control image is taken from an identical assay without primary antibody.

    For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

    *Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

  • All lanes : No Primary Antibody

    Lane 1 : Liver (Human) Tissue Lysate
    Lane 2 : Liver (Mouse) Tissue Lysate
    Lane 3 : Liver (Rat) Tissue Lysate
    Lane 4 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate
    Lane 5 : NIH 3T3 (Mouse embryonic fibroblast cell line) Whole Cell Lysate
    Lane 6 : PC12 (Rat adrenal pheochromocytoma cell line) Whole Cell Lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : Donkey Anti-Mouse IgG H&L (HRP) (ab205724) at 1/2000 dilution

    Performed under reducing conditions.

    Exposure time: 15 seconds


    This blot was produced using a 4-12% Bis-tris gel under the MOPS buffer system. The gel was run at 200V for 50 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was incubated overnight with 2% Bovine Serum Albumin at 4°C. Any non-specific background binding was assessed by incubating the membrane with only the secondary antibody (ab205724), and visualised using ECL development solution ab133406.

  • All lanes : Anti-beta Actin antibody [mAbcam 8226] - Loading Control (ab8226) at 1 µg/ml

    Lane 1 : Liver (Human) Tissue Lysate
    Lane 2 : Liver (Mouse) Tissue Lysate
    Lane 3 : Liver (Rat) Tissue Lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : ab205724 (Left Image) at 1/2000 and a competitor secondary (Right Image) at 1/2000. Notice the increased background of the competitor product.

    Performed under reducing conditions.

    Observed band size: 42 kDa why is the actual band size different from the predicted?


    Exposure time: 15 seconds


    This blot was produced using a 4-12% Bis-tris gel under the MOPS buffer system. The gel was run at 200V for 50 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour using 2% Bovine Serum Albumin before being incubated with ab8226 overnight at 4°C. Antibody binding was detected using ab205724 (Left Image) and a competitor secondary (Right Image), and visualised using ECL development solution ab133406.

  • All lanes : No Primary Antibody

    Lane 1 : Liver (Human) Tissue Lysate
    Lane 2 : Liver (Mouse) Tissue Lysate
    Lane 3 : Liver (Rat) Tissue Lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    All lanes : ab205724 (Left Image) 1/2000 and a competitor secondary (Right Image) 1/2000. Notice the increased background of the competitor product.

    Performed under reducing conditions.

    Exposure time: 15 seconds


    This blot was produced using a 4-12% Bis-tris gel under the MOPS buffer system. The gel was run at 200V for 50 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was incubated overnight with 2% Bovine Serum Albumin at 4°C. Any non-specific background binding was assessed by incubating the membrane with ab205724 (Left Image) and a competitor secondary (Right Image), and visualised using ECL development solution ab133406.

References

This product has been referenced in:
  • Maggi EC  et al. Retinoblastoma-binding protein 2 (RBP2) is frequently expressed in neuroendocrine tumors and promotes the neoplastic phenotype. Oncogenesis 5:e257 (2016). Read more (PubMed: 27548814) »
See 1 Publication for this product

Customer reviews and Q&As

1-2 of 2 Abreviews or Q&A

GAPDH western

Excellent
Abreviews
Application
Western blot
5ug liver lysate (abcam ab29889) 10ug whole cell lysate (human ES cells)
Blocking buffer - 5%BSA/1%OvAlb/PBS (also works with 5% milk powder) for 3hours
Primary antibody - GAPDH (1:10000 expected size 37kDa) 14hours at 4C
Secondary diluted in TBST (1:5000) for 3 hours at room temperature 3hours at room temperature
ECL prime
5 second exposure
Lane 1. Liver lysate
Lane 2. Ladder
Lane 3. Human ES cell lysate

Abcam user community

Verified customer

Submitted Mar 24 2016

Application
Western blot
Excellent specificity.
Good sensitivity.
WB conditions:
- Cell lisates: 20 ug protein
- non-reducing
- Primary antibody: Mouse anti-GRP78, clon 40/BiP, Cat. No. 610979. BD Biosciences. Dilution 1:1000
- Secondary antibody: 1:6000
- Developer: ECL Prime, Amersham, GE Helathcare Life Sciences, Product code RPN2232
- Expusure time: 1 second, 3th film out 4

Abcam user community

Verified customer

Submitted Feb 01 2016

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