概述

  • 产品名称Anti-NOTCH3抗体
    参阅全部 NOTCH3 一抗
  • 描述
    兔多克隆抗体to NOTCH3
  • 经测试应用适用于: WB, IHC-P, ICC/IF, IHC-FoFr, IHC-Fr, ICCmore details
  • 种属反应性
    与反应: Mouse, Rat, Human
  • 免疫原

    Synthetic peptide corresponding to Human NOTCH3 aa 2300 to the C-terminus (C terminal) conjugated to Keyhole Limpet Haemocyanin (KLH).
    (Peptide available as ab26878)

  • 阳性对照
    • This antibody gave a positive signal in the following Human Cell lysates: K562 and CACO-2

性能

应用

Our Abpromise guarantee covers the use of ab23426 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
WB Use a concentration of 1 µg/ml. Detects a band of approximately 97,280 kDa (predicted molecular weight: 244 kDa).
IHC-P Use a concentration of 5 µg/ml. Perform heat mediated antigen retrieval before commencing with IHC staining protocol. PubMed: 19965627
ICC/IF Use at an assay dependent concentration. PubMed: 20680961
IHC-FoFr Use at an assay dependent concentration.
IHC-Fr 1/300.
ICC Use at an assay dependent concentration.

靶标

  • 功能Functions as a receptor for membrane-bound ligands Jagged1, Jagged2 and Delta1 to regulate cell-fate determination. Upon ligand activation through the released notch intracellular domain (NICD) it forms a transcriptional activator complex with RBPJ/RBPSUH and activates genes of the enhancer of split locus. Affects the implementation of differentiation, proliferation and apoptotic programs.
  • 组织特异性Ubiquitously expressed in fetal and adult tissues.
  • 疾病相关Defects in NOTCH3 are the cause of cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) [MIM:125310]. CADASIL causes a type of stroke and dementia of which key features include recurrent subcortical ischemic events and vascular dementia. The disorder affects relatively young adults of both sexes. Mutations affect highly conserved cysteine residues within epidermal growth factor (EGF)-like repeat domains in the extracellular part of the receptor.
  • 序列相似性Belongs to the NOTCH family.
    Contains 5 ANK repeats.
    Contains 34 EGF-like domains.
    Contains 3 LNR (Lin/Notch) repeats.
  • 翻译后修饰Synthesized in the endoplasmic reticulum as an inactive form which is proteolytically cleaved by a furin-like convertase in the trans-Golgi network before it reaches the plasma membrane to yield an active, ligand-accessible form. Cleavage results in a C-terminal fragment N(TM) and a N-terminal fragment N(EC). Following ligand binding, it is cleaved by TNF-alpha converting enzyme (TACE) to yield a membrane-associated intermediate fragment called notch extracellular truncation (NEXT). This fragment is then cleaved by presenilin dependent gamma-secretase to release a notch-derived peptide containing the intracellular domain (NICD) from the membrane.
    Phosphorylated.
  • 细胞定位Cell membrane and Nucleus. Following proteolytical processing NICD is translocated to the nucleus.
  • Information by UniProt
  • 数据库链接
  • 别名
    • CADASIL antibody
    • CASIL antibody
    • NOTC3_HUMAN antibody
    • Notch 3 antibody
    • Notch 3 intracellular domain antibody
    • Notch homolog 3 antibody
    • Notch3 antibody
    see all

Anti-NOTCH3 antibody 图像

  • All lanes : Anti-NOTCH3 antibody (ab23426) at 1 µg/ml

    Lane 1 : K562 whole cell lysate (ab29306)
    Lane 2 : Caco 2 whole cell lysate (ab3950)
    Lane 3 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    Goat polyclonal to Rabbit IgG - H&L (HRP) at 1/50000 dilution
    Developed using the ECL technique

    Performed under reducing conditions.

    Predicted band size : 244 kDa
    Observed band size : 97,280 kDa (why is the actual band size different from the predicted?)
    Additional bands at : 105 kDa,30 kDa. We are unsure as to the identity of these extra bands.

    Exposure time : 30 seconds

    This blot was produced using a 3-8% Tris Acetate gel under the TA buffer system. The gel was run at 150V for 60 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour using 5% Bovine Serum Albumin before being incubated with ab23426 overnight at 4°C. Antibody binding was detected using an anti-rabbit antibody conjugated to HRP, and visualised using ECL development solution.

    The band observed at 97 kDa is thought to correspond to the notch-derived peptide containing the intracellular domain (NICD) of NOTCH3 as described in the literature (PMID:10712431).

  • ab23426 staining NOTCH3 in mouse embryo tissue (E16.5) by Immunohistochemistry (Formalin/PFA-fixed paraffin embedded sections). Tissue underwent fixation in formaldehyde, heat-mediated antigen retreival in citrate buffer pH6.0 and blocking for 15 minutes at 10°C (5 minutes/peroxidase blocking and 10 minutes/protein blocking). The primary antibody was diluted 1/250 and incubated with sample for 15 minutes at 20°C. A HRP-conjugated goat polyclonal to rabbit IgG was used undiluted as the secondary.

    See Abreview

  • ab23426 staining NOTCH3 in human breast cancer tissue sections by immunohistochemistry (Formalin/PFA-fixed paraffin embedded sections). Tissue underwent fixation in paraformaldehyde, heat-mediated antigen retrieval in citrate buffer pH6.0 and blocking for 15 minutes at 20°C (5 minutes for peroxidase blocking and 10 minutes for protein blocks). The primary antibody was diluted 1/250 and incubated with sample for 45 minutes at 20°C. A HRP-conjugated goat polyclonal to rabbit IgG was used undiluted as secondary.

    See Abreview

  • IHC-FoFr image of NOTCH3 staining in Rat cerebral cortex (left) and the paraventricular hypothalamic nucleus (right) sections. The sections used came from animals perfused fixed with Paraformaldehyde 4%, in phosphate buffer 0.2M. Following postfixation in the same fixative overnight, the brains were cryoprotected in sucrose 30% overnight. Immunostainings were performed using the ‘free floating’ technique.

    See Abreview

  • IHC image of NOTCH3 staining in human breast carcinoma formalin fixed paraffin embedded tissue section, performed on a Leica BondTM 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 ab23426, 5µg/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.
  • ICC/IF image of ab23426 stained mouse embryonic stem 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 (ab23426, 5µg/ml) overnight at +4°C. The secondary antibody (green) was ab96899, DyLight® 488 goat anti-rabbit IgG (H+L) 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.
  • ICC/IF image of ab23426 stained MCF7 cells. The cells were 100% methanol fixed (5 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 (ab23426, 5µg/ml) overnight at +4°C. The secondary antibody (green) was ab96899, DyLight® 488 goat anti-rabbit IgG (H+L) 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.

  • ab14404 staining NOTCH3 in the COS1 fibroblast cell line from Monkey Kkidney by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with paraformaldehyde, permeabilized with Triton X-100 0.1% in PBS and blocked with 1% BSA for 30 minutes at 25°C. Samples were incubated with primary antibody (1/200) for 16 hour at 4°C. An Alexa Fluor®488-conjugated Goat anti-rabbit polyclonal (1/500) was used as the secondary antibody.

    See Abreview

Anti-NOTCH3 antibody (ab23426)参考文献

This product has been referenced in:
  • Tripathi R  et al. Clinical impact of de-regulated Notch-1 and Notch-3 in the development and progression of HPV-associated different histological subtypes of precancerous and cancerous lesions of human uterine cervix. PLoS One 9:e98642 (2014). IHC-P ; Human . Read more (PubMed: 24905589) »
  • Murta D  et al. In vivo notch signaling blockade induces abnormal spermatogenesis in the mouse. PLoS One 9:e113365 (2014). IHC ; Mouse . Read more (PubMed: 25412258) »

See all 14 Publications for this product

Product Wall

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunocytochemistry/ Immunofluorescence
Sample Mouse Cell (NIH3T3)
Specification NIH3T3
Fixative Paraformaldehyde
Permeabilization Yes - 0.1% Triton
Blocking step Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 24°C
Username

Abcam user community

Verified customer

提交于 Mar 19 2013

Application Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Antigen retrieval step Heat mediated - Buffer/Enzyme Used: Citrate Buffer pH6.0
Sample Human Tissue sections (Human melanoma xenograft in mouse)
Specification Human melanoma xenograft in mouse
Blocking step Dakocytomation X0909 as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 100% · Temperature: RT°C
Fixative Formaldehyde
Username

Hongwei Shao

Verified customer

提交于 May 08 2014


Unfortunately it is not possible for us to sell the pre-immune serum of our rabbits.
I am sorry that on this occasion I could not be of more help.

Application Immunocytochemistry/ Immunofluorescence
Sample Monkey Cell (COS1 cell line)
Specification COS1 cell line
Fixative Paraformaldehyde
Permeabilization Yes - 0.1% Triton X100
Blocking step BSA as blocking agent for 30 minute(s) · Concentration: 1% · Temperature: 25°C
Username

Abcam user community

Verified customer

提交于 Feb 11 2013

Thank you for contacting us.
I am pleased to hear that you consider participating in our AbTrial programme. For the reason that the AbTrial Code is restricted for 4 month after issuing I would suggest you let me know in January if you decide to ac...

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Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunocytochemistry/ Immunofluorescence
Sample Human Cell (HUVEK cells)
Specification HUVEK cells
Fixative Paraformaldehyde
Permeabilization Yes - 0.1% Triton X100
Blocking step BSA as blocking agent for 30 minute(s) · Concentration: 1% · Temperature: 25°C
Username

Abcam user community

Verified customer

提交于 Nov 23 2012

Thank you for contacting Abcam regarding ab23426.


Regarding the immunogen information, please contact us directly as the specific sequence is proprietary.


The entire sequence is contained within the intracellular domain....

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Thank you for your reply and for the additional information.

The antibody will be less stable at 1:100, though the storage at -20C will help prevent degradation. I think it's possible that the antibody is still fine to use, though I would rec...

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Thank you for contacting us with your question.

The antibody stock solution is provided in 1% BSA, howeverit is fine tomake your working dilutions ina buffer appropriate for your assay (such as PBS, PBST, TBS, or TBST)with or without BSA. We ...

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Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunohistochemistry (Frozen sections)
Sample Human Tissue sections (endometrial cancer)
Specification endometrial cancer
Fixative Acetone
Permeabilization No
Blocking step Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 20°C
Username

Abcam user community

Verified customer

提交于 Mar 21 2012

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