概述

  • 产品名称
    Anti-PIP2抗体[2C11]
    参阅全部 PIP2 一抗
  • 描述
    小鼠单克隆抗体[2C11] to PIP2
  • 特异性
    Reacts with PtdIns(4,5)P2, Ptd(4)P and Ptd(3,4,5)P3.
  • 经测试应用
    适用于: IHC-P, ICC/IF, ELISA, Neutralising, IPmore details
  • 种属反应性
    与反应: Mouse, Rat, Human
  • 免疫原

    Liposomes (prepared in PBS with lipid A, phosphatidylcholine (PC) and cholesterol) containing synthetic di-palmitoyl PtdIns(4,5)P2.

  • 阳性对照
    • IHC-P: FFPE human kidney normal and FFPE mouse normal brain. ICC: HepG2 cell line, Neuro2a cell line
  • 常规说明

    This antibody clone is manufactured by Abcam.

    For testing in lipid dot blot assay, follow the protocol used in Thomas et al. Biochem Soc Trans 27:648-52 (1999) (PMID: 10917659, please see the References tab).

    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.

性能

应用

Our Abpromise guarantee covers the use of ab11039 in the following tested applications.

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

应用 Ab评论 说明
IHC-P Use a concentration of 1 - 5 µg/ml. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
ICC/IF Use a concentration of 1 - 5 µg/ml.
ELISA Use at an assay dependent concentration.
Neutralising Use at an assay dependent concentration.
IP Use at an assay dependent concentration.

靶标

  • 相关性
    Phosphatidylinositol 4,5-biphosphate (PIP2) is a membrane phospholipid that has been implicated in a variety of cellular processes, including synaptic vesicle recycling and signal transduction pathways. PLCD4 hydrolyzes PIP2 to generate 2 second messenger molecules diacylglycerol (DAG) and inositol 1,4,5-trisphosphate (IP3).

图片

  • ICC/IF image of ab11039 stained human 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 (ab11039, 5 µg/ml) overnight at +4°C.

    The secondary antibody (green) was Alexa Fluor® 488 goat anti-mouse IgM (H+L) used at a 1/1000 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). This antibody also gave a positive IF result in HeLa, Hek293 and MCF7 cells.

  • IHC image of PIP2 staining in mouse normal brain formalin fixed paraffin embedded tissue section. The section was pre-treated using pressure cooker heat mediated antigen retrieval with sodium citrate buffer (pH6) for 30mins. The section was incubated with ab11039, 1µg/ml overnight at +4°C. An HRP-conjugated secondary (Ab98679, 1/1000 dilution) was used for 1hr at room temperature. The section was counterstained with haematoxylin and mounted with DPX.

  • Immunofluorescence analysis of Human SaOS-2 (Human osteosarcoma) cells, staining PIP2 with ab11039. Cells were either unstimulated (upper panel) or stimulated with direct current (lower panel).

    Cells were fixed in formaldehyde, permeabilized and then blocked with 1% BSA for 20 min. Cells were then incubated with a primary antibody (1/200) overnight at 4°C. A FITC-conjugated anti-mouse IgG was used as the secondary antibody.
  • IHC image of PIP2 staining in a formalin fixed, paraffin embedded human normal kidney tissue section*, performed on a Leica Bond™ system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab11039, 1µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

     

    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

文献

This product has been referenced in:

See all 12 Publications for this product

客户评价及客户问答

We do share your concerns regarding mouse on mouse staining of kidney tissues. At Abcam we recommend following the attached mouse on mouse protocol tips to reduce background from this type of staining. While we do not have data regarding kidney tiss...

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Abcam has not validated the combination of species/application used in this Abreview.
Application
Western blot
Sample
Human Cell lysate - whole cell (HEK293)
Loading amount
20 µg
Specification
HEK293
Gel Running Conditions
Non-reduced Denaturing (10% gel)
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 21°C
Username

Abcam user community

Verified customer

提交于 Feb 01 2013

Thank you for taking the time to complete our questionnaire and contact us. I am sorry to hear you have had difficulty obtaining satisfactory results from this antibody.

The details you have kindly provided will enable us to investigate this ...

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Thank you for taking the time to contact us. I am sorry to hear you have had difficulty obtaining satisfactory results from these antibodies.

I would like to reassure you that this antibodies are tested and covered by our 6 month guarantee for...

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Thank you for contacting us.

I personally don't have experience with using a freeze-thaw permeabilization, but from doing a quick literature search it does seem to be a good method for staining intracellular membrane proteins.

http...

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Thank you very much for your reply.
Since PIP2 seems to be ubiquitously expressed, I would expect most, if not all, cells to be positive controls. For ab11039, HepG2 may be the best positive control since the antibody has been tested in this cell ...

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Thank you again for your reply and for your patience.
I have followed-up with the lab and the secondary antibody mentioned in the image caption was actually an anti-mouse IgM, not IgG. We have corrected this in the caption. Thank you very much for...

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Thank you very much for your reply.
I'll need to check with the lab to confirm the type of secondary antibody that was used in this ICC/IF protocol. The anti-mouse IgG secondary antibody described in the image caption would still probably cross-re...

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Thank you very much for contacting us with your question.
I've looked through some literature, and it looks like PIP2 can be found in the nucleus as well as on the plasma membrane-
http://jcs.biologists.org/content/114/13/2501.long
http:...

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Thank you for your call today and for letting us know about the trouble with this antibody.

Your credit note ID is ***

I am sorry that this antibody did not perform as stated on the datasheet. If payment has already been made on the ...

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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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