Anti-M6PR (cation independent) antibody [2G11] (ab2733)

Overview

  • Product nameAnti-M6PR (cation independent) antibody [2G11]
    See all M6PR (cation independent) primary antibodies
  • Description
    Mouse monoclonal [2G11] to M6PR (cation independent)
  • Tested applicationsSuitable for: Flow Cyt, ICC/IF, IHC-FoFr, WB, IPmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Cow, Human, African Green Monkey
    Predicted to work with: Non Human PrimatesDoes not react with: Hamster
  • Immunogen

    Purified bovine 300 kDa Mannose 6 Phosphate Receptor (Cation independent).

  • EpitopeThis antibody is shown to recognize an epitope in the extracellular domain of Mannose 6 Phosphate Receptor.
  • Positive control
    • This antibody gave a positive signal in the following whole cell lysates: HeLa, SHSY-5Y, HepG2, MEF1. In Flow Cytometry, this antibody gave a positive signal in A431 cells.
  • General notes

    Alternative versions available:

    Anti-M6PR (cation independent) antibody (Alexa Fluor® 488) [2G11] (ab205812)
    Anti-M6PR (cation independent) antibody (Alexa Fluor® 647) [2G11] (ab205813)

Properties

Applications

Our Abpromise guarantee covers the use of ab2733 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
Flow Cyt Use 1µg for 106 cells.
ICC/IF Use a concentration of 5 - 10 µg/ml.
IHC-FoFr Use at an assay dependent concentration.
WB Use a concentration of 5 - 10 µg/ml. Detects a band of approximately 300 kDa (predicted molecular weight: 300 kDa). Abcam recommends using BSA as the blocking agent.
IP Use at an assay dependent concentration.

Target

  • FunctionTransport of phosphorylated lysosomal enzymes from the Golgi complex and the cell surface to lysosomes. Lysosomal enzymes bearing phosphomannosyl residues bind specifically to mannose-6-phosphate receptors in the Golgi apparatus and the resulting receptor-ligand complex is transported to an acidic prelyosomal compartment where the low pH mediates the dissociation of the complex. This receptor also binds IGF2. Acts as a positive regulator of T-cell coactivation, by binding DPP4.
  • Sequence similaritiesBelongs to the MRL1/IGF2R family.
    Contains 1 fibronectin type-II domain.
  • DomainContains 15 repeating units of approximately 147 AA harboring four disulfide bonds each. The most highly conserved region within the repeat consists of a stretch of 13 AA that contains cysteines at both ends.
  • Cellular localizationLysosome membrane. Colocalized with DPP4 in internalized cytoplasmic vesicles adjacent to the cell surface.
  • Information by UniProt
  • Database links
  • Alternative names
    • 300 kDa mannose 6 phosphate receptor antibody
    • 300 kDa mannose 6-phosphate receptor antibody
    • Cation independent mannose 6 phosphate receptor antibody
    • Cation-independent mannose-6-phosphate receptor antibody
    • CD222 antibody
    • CD222 antigen antibody
    • CI Man 6 P receptor antibody
    • CI Man-6-P receptor antibody
    • CI MPR antibody
    • CI-M6PR antibody
    • CI-MPR antibody
    • CIMPR antibody
    • IGF 2 receptor antibody
    • IGF 2R antibody
    • IGF II receptor antibody
    • IGF-II receptor antibody
    • IGF2 receptor antibody
    • Igf2r antibody
    • Insulin like growth factor 2 receptor antibody
    • Insulin like growth factor II receptor antibody
    • Insulin-like growth factor 2 receptor antibody
    • Insulin-like growth factor II receptor antibody
    • M6P R antibody
    • M6P/IGF2 receptor antibody
    • M6P/IGF2R antibody
    • M6PR antibody
    • mannose 6 phosphate receptor antibody
    • mannose 6 phosphate receptor, cation independent antibody
    • MPR 300 antibody
    • MPR300 antibody
    • MPRI antibody
    • MPRI_HUMAN antibody
    see all

Anti-M6PR (cation independent) antibody [2G11] images

  • ICC/IF image of ab2733 stained human HeLa cells. The cells were methanol fixed (5 min) and incubated with the antibody (ab2733, 1µg/ml) for 1h at room temperature. The secondary antibody (green) was Alexa Fluor® 488 goat anti-mouse IgG (H+L) used at a 1/1000 dilution for 1h. Image-iTTM FX Signal Enhancer was used as the primary blocking agent, 5% BSA (in TBS-T) was used for all other blocking steps. DAPI was used to stain the cell nuclei (blue). Alexa Fluor® 594 WGA was used to label plasma membranes (red).

  • Immunofluorescent imaging of human cells (U2OS) with ab2733 confirms the specificity of this antibody, with the expected perinuclear vesicular staining of lysosomes. 

    IF was performed with a standard paraformaldehyde technique (fixed in PBS buffered PFH 4% for 5 minutes, permeabilised with 0.5% triton-PBS for 5 minutes, blocked with 5% milk / 0.2% tween for one hour.  Primary antibody used at 1/100 in 5% milk / 0.2% TWEEN for one hour, secondary antibody for 30 minutes.  All blocking and incubation steps carried out at 37 degrees.

  • Immunostained frozen sections showing that mannose 6 phosphate receptor is localised in the outermost epithelial cell layer.

    This picture was kindly supplied as part of the review submitted by Marko Nykanen.

  • ab2733 positively staining formaldehyde fixed Human HEK 293 cells (red) in conjunction with goat anti mouse (Alexa 546). Nuclear staining was obtained using Hoechst.
    This image is an edited version of an image received courtesy of an Abreview submitted by Kun Liu on 19 September 2005. We do not have any further information relating to this image.

    See Abreview

  • ICC/IF image of ab2733 stained HepG2 cells. The cells were 4% PFA fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab2733, 10µg/ml) overnight at +4°C. The secondary antibody (green) was DyLight® 488 goat anti-mouse IgG - H&L, pre-adsorbed (ab96879) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.

  • Overlay histogram showing HeLa cells stained with ab2733 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab2733, 1µg/1x106 cells) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was mouse IgG2a [ICIGG2A] (ab91361, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed.
  • Lanes 1 - 4 : Anti-M6PR (cation independent) antibody [2G11] (ab2733) at 5 µg/ml
    Lanes 5 - 8 : Anti-M6PR (cation independent) antibody [2G11] (ab2733) at 10 µg/ml

    Lane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate
    Lane 2 : SHSY-5Y (Human neuroblastoma cell line) Whole Cell Lysate
    Lane 3 : HepG2 (Human hepatocellular liver carcinoma cell line) Whole Cell Lysate
    Lane 4 : MEF1 (Mouse embryonic fibroblast cell line) Whole Cell Lysate
    Lane 5 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate
    Lane 6 : SHSY-5Y (Human neuroblastoma cell line) Whole Cell Lysate
    Lane 7 : HepG2 (Human hepatocellular liver carcinoma cell line) Whole Cell Lysate
    Lane 8 : MEF1 (Mouse embryonic fibroblast cell line) Whole Cell Lysate

    Lysates/proteins at 40 µg per lane.

    Secondary
    Goat Anti-Mouse IgG H&L (HRP) preadsorbed (ab97040) at 1/5000 dilution
    Developed using the ECL technique

    Performed under reducing conditions.

    Predicted band size : 300 kDa
    Observed band size : 300 kDa


    Exposure time : 1 minute

References for Anti-M6PR (cation independent) antibody [2G11] (ab2733)

This product has been referenced in:
  • Fuse A  et al. VPS29-VPS35 intermediate of retromer is stable and may be involved in the retromer complex assembly process. FEBS Lett 589:1430-6 (2015). Read more (PubMed: 25937119) »
  • Munson MJ  et al. mTOR activates the VPS34-UVRAG complex to regulate autolysosomal tubulation and cell survival. EMBO J 34:2272-90 (2015). Read more (PubMed: 26139536) »

See all 44 Publications for this product

Product Wall

Thank you for confirming these details and for your cooperation. The details provided enable us to closely monitor the quality of our products.

I am sorry this product did not perform as stated on the datasheet and for the inconvenience this h...

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Thank you, I received the data this time. For a protocol modification, I suggest trying preparing a sample by heating at 60C for 10 minutes instead of 95 for 5 minutes to minimize aggregation of the trans-membrane domains. It is strange that both antib...

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Thank you for bringing this to our attention. I will need a few more details of your protocol. I was not able to open the file you sent.

Can you please tell me how you prepared your lysates (did you use protease inhibitors), and how you heat...

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Thank you for contacting us regarding ab2733. I have confirmed with the laboratory that this particular antibody is only batch tested in IF, but has been tested in WB. I have attached an image for you. We have recently done some optimisation ...

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I'm glad that you have been seeing better results with blocking with BSA. If you are now having difficulty with non-specific bands, increasing the dilution of the primary and secondary antibody as well as adding 1 % BSA and 0.2% Tween 20 into the dilue...

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Thank you for your reply. I am sorry this product did not perform as stated on the datasheet and for the inconvenience this has caused. As requested, I have issued a free of charge replacement for ab8093. To check the status of the order please...

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Thank you for your reply. As this antibody, ab2733 is a monoclonal antibody I would not expect much lot to lot variation.  It is possible that there was something wrong with just this particular vial as we have not received reports of other problem...

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Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunocytochemistry/ Immunofluorescence
Sample Cow Cell (MDBK - Madin-Darby Bovine Kidney)
Specification MDBK - Madin-Darby Bovine Kidney
Fixative Paraformaldehyde
Permeabilization Yes - 0.5% Triton X-100
Blocking step Serum as blocking agent for 20 minute(s) · Concentration: 10% · Temperature: 25°C
Username

Abcam user community

Verified customer

Submitted Jun 13 2011

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunocytochemistry/ Immunofluorescence
Sample Human Cell (HEK293)
Specification HEK293
Fixative 100% EtOH
Permeabilization Yes - 0.1% Triton X-100
Blocking step Gelatin as blocking agent for 40 minute(s) · Concentration: 0.1% · Temperature: 25°C
Username

Abcam user community

Verified customer

Submitted Jan 14 2011

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunocytochemistry/ Immunofluorescence
Sample Human Cell (HeLa cells)
Specification HeLa cells
Fixative Paraformaldehyde
Permeabilization Yes - 0.1% Triton X-100
Blocking step Serum as blocking agent for 10 minute(s) · Concentration: 10% · Temperature: 20°C
Username

Dr. Margaret Robinson

Verified customer

Submitted Dec 02 2010

1-10 of 31 Abreviews or Q&A

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