Recombinant Anti-PODXL antibody [EPR9518] (ab150358)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR9518] to PODXL
- Suitable for: Flow Cyt (Intra), WB, IHC-P, ICC/IF
- Knockout validated
- Reacts with: Human
Related conjugates and formulations
Overview
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Product name
Anti-PODXL antibody [EPR9518]
See all PODXL primary antibodies -
Description
Rabbit monoclonal [EPR9518] to PODXL -
Host species
Rabbit -
Tested applications
Suitable for: Flow Cyt (Intra), WB, IHC-P, ICC/IFmore details
Unsuitable for: IP -
Species reactivity
Reacts with: Human -
Immunogen
Recombinant fragment within Human PODXL aa 300-500. The exact sequence is proprietary.
Database link: O00592 -
Positive control
- WB: Raji, HeLa, HCT116 and HAP1 whole cell lysate. Human fetal kidney lysate. IHC-P: Human kidney tissue. Human hepatocellular carcinoma, breast carcinoma and endometrial carcinoma tissue. Human glioma tissue. ICC/IF: HeLa cells. Flow Cyt (intra): HeLa cells.
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General notes
This product is a recombinant monoclonal antibody, which offers several advantages including:
- - High batch-to-batch consistency and reproducibility
- - Improved sensitivity and specificity
- - Long-term security of supply
- - Animal-free production
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.
We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.
Mouse, Rat: We have preliminary internal testing data to indicate this antibody may not react with these species. Please contact us for more information.
Properties
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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. -
Storage buffer
Preservative: 0.01% Sodium azide
Constituents: 0.05% BSA, 40% Glycerol, PBS -
Concentration information loading...
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Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
EPR9518 -
Isotype
IgG -
Research areas
Associated products
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Alternative Versions
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Compatible Secondaries
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Isotype control
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KO cell lines
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KO cell lysates
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Positive Controls
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Recombinant Protein
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab150358 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 |
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Flow Cyt (Intra) |
1/250.
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WB |
1/1000 - 1/10000. Detects a band of approximately 165 kDa (predicted molecular weight: 58 kDa).
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IHC-P | (2) |
1/1000. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
For unpurified use at 1/250 - 1/500. |
ICC/IF | (1) |
1/100.
For unpurified use at 1/500. |
Notes |
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Flow Cyt (Intra)
1/250. |
WB
1/1000 - 1/10000. Detects a band of approximately 165 kDa (predicted molecular weight: 58 kDa). |
IHC-P
1/1000. Perform heat mediated antigen retrieval before commencing with IHC staining protocol. For unpurified use at 1/250 - 1/500. |
ICC/IF
1/100. For unpurified use at 1/500. |
Target
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Function
Involved in the regulation of both adhesion and cell morphology and cancer progression. Function as an anti-adhesive molecule that maintains an open filtration pathway between neighboring foot processes in the podocyte by charge repulsion. Acts as a pro-adhesive molecule, enhancing the adherence of cells to immobilized ligands, increasing the rate of migration and cell-cell contacts in an integrin-dependent manner. Induces the formation of apical actin-dependent microvilli. Involved in the formation of a preapical plasma membrane subdomain to set up inital epithelial polarization and the apical lumen formation during renal tubulogenesis. Plays a role in cancer development and aggressiveness by inducing cell migration and invasion through its interaction with the actin-binding protein EZR. Affects EZR-dependent signaling events, leading to increased activities of the MAPK and PI3K pathways in cancer cells. -
Tissue specificity
Glomerular epithelium cell (podocyte). -
Sequence similarities
Belongs to the podocalyxin family. -
Domain
Both the O-glycan-rich domain of the extracellular domain and the C-terminus PDZ-binding motif (DTHL) in the cytoplasmic tail harbor an apical sorting signal. The cytoplasmic domain is necessary for the apical membrane targeting and renal tubulogenesis. The cytoplasmic C-terminus PDZ-binding motif (DTHL) is essential for interaction with SLC9A3R1 and for targeting SLC9A3R1 to the apical cell membrane. The extracellular domain is necessary for microvillus formation (By similarity). The large highly anionic extracellular domain allows to maintain open filtration pathways between neighboring podocyte foot processes. -
Post-translational
modificationsN- and O-linked glycosylated. Sialoglycoprotein. -
Cellular localization
Apical cell membrane. Cell projection, lamellipodium. Cell projection, filopodium. Cell projection, ruffle. Cell projection, microvillus. Membrane raft. Membrane. In single attached epithelial cells is restricted to a preapical pole on the free plasma membrane whereas other apical and basolateral proteins are not yet polarized. Colocalizes with SLC9A3R2 at the apical plasma membrane during epithelial polarization. Colocalizes with SLC9A3R1 at the trans-Golgi network (transiently) and at the apical plasma membrane. Its association with the membrane raft is transient. Colocalizes with actin filaments, EZR and SLC9A3R1 in a punctate pattern at the apical cell surface where microvilli form. Colocalizes with EZR and SLC9A3R2 at the apical cell membrane of glomerular epithelium cells (By similarity). Forms granular, punctuated pattern, forming patches, preferentially adopting a polar distribution, located on the migrating poles of the cell or forming clusters along the terminal ends of filipodia establishing contact with the endothelial cells. Colocalizes with the submembrane actin of lamellipodia, particularly associated with ruffles. Colocalizes with vinculin at protrusions of cells. Colocalizes with ITGB1. Colocalizes with PARD3, PRKCI, EXOC5, OCLN, RAB11A and RAB8A in apical membrane initiation sites (AMIS) during the generation of apical surface and luminogenesis (By similarity). - Information by UniProt
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Database links
- Entrez Gene: 5420 Human
- Omim: 602632 Human
- SwissProt: O00592 Human
- Unigene: 744213 Human
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Form
There are 2 isoforms produced by alternative splicing. -
Alternative names
- GCTM-2 antigen antibody
- Gp2 antibody
- Gp200 antibody
see all
Images
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All lanes : Anti-PODXL antibody [EPR9518] (ab150358) at 1/10000 dilution
Lane 1 : Wild-type HeLa cell lysate
Lane 2 : PODXL knockout HeLa cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 58 kDa
Observed band size: 160 kDa why is the actual band size different from the predicted?Lanes 1- 2: Merged signal (red and green). Green - ab150358 observed at 160 kDa. Red - Anti-GAPDH antibody [6C5] - Loading Control (ab8245) observed at 37 kDa.
ab150358 was shown to react with PODXL in wild-type HeLa cells in western blot. Loss of signal was observed when knockout cell line ab264984 (knockout cell lysate ab257210) was used. Wild-type HeLa and PODXL knockout HeLa cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab150358 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) overnight at 4°C at a 1 in 10000 dilution and a 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye®800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye®680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PODXL antibody [EPR9518] (ab150358)
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human kidney tissue sections labeling PODXL with purified ab150358 at 1/1000 dilution (0.44 μg/ml). Heat mediated antigen retrieval was performed using ab93684 (Tris/EDTA buffer, pH 9.0). Tissue was counterstained with hematoxylin. ImmunoHistoProbe one step HRP Polymer (ready to use) is the secondary antibody.
PBS instead of the primary antibody was used as the negative control.
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All lanes : Anti-PODXL antibody [EPR9518] (ab150358) at 1/1000 dilution
Lane 1 : Wild-type HCT116 cell lysate
Lane 2 : PODXL knockout HCT116 cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 58 kDa
Observed band size: 200 kDa why is the actual band size different from the predicted?Lanes 1- 2: Merged signal (red and green). Green - ab150358 observed at 200 kDa. Red - Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) observed at 50 kDa.
ab150358 was shown to react with PODXL in wild-type HCT116 cells in western blot. Loss of signal was observed when knockout cell line ab266887 (knockout cell lysate ab257211) was used. Wild-Type HCT116 and PODXL knockout HCT116 cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab150358 and Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) overnight at 4°C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye®800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye®680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
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All lanes : Anti-PODXL antibody [EPR9518] (ab150358) at 1/1000 dilution
Lane 1 : Wild-type HAP1 whole cell lysate
Lane 2 : PODXL knockout HAP1 whole cell lysate
Lane 3 : Raji whole cell lysate
Lane 4 : HeLa whole cell lysate
Lysates/proteins at 20 µg per lane.
Predicted band size: 58 kDaLanes 1 - 4: Merged signal (red and green). Green - ab150358 observed at 160 kDa. Red - loading control, ab9484, observed at 37 kDa.
ab150358 was shown to specifically react with PODXL in wild-type cells as signal was lost in PODXL knockout cells. Wild-type and PODXL knockout samples were subjected to SDS-PAGE. Ab150358 and ab9484 (Mouse anti-GAPDH loading control) were incubated overnight at 4°C at a 1/1000 dilution and 1/20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed ab216776 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.
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Anti-PODXL antibody [EPR9518] (ab150358) at 1/10000 dilution (purified) + Human fetal kidney lysates at 15 µg
Secondary
Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/2000 dilution
Predicted band size: 58 kDa
Observed band size: 165 kDa why is the actual band size different from the predicted?Blocking and diluting buffer: 5% NFDM/TBST.
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Immunocytochemistry/Immunofluorescence analysis of HeLa (human cervix adenocarcinoma) cells labeling PODXL with purified ab150358 at 1/500. Cells were fixed with 100% methanol. An Alexa Fluor® 488-conjugated goat anti-rabbit IgG (ab150077) at 1/1000 dilution was used as the secondary antibody. Nuclei couterstained with DAPI (blue).
Secondary Only Control: PBS was used instead of the primary antibody as the negative control.
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All lanes : Anti-PODXL antibody [EPR9518] (ab150358) at 1/1000 dilution (unpurified)
Lane 1 : Raji lysate
Lane 2 : HeLa lysate
Lane 3 : Human fetal kidney lysate
Lysates/proteins at 10 µg per lane.
Secondary
All lanes : HRP labelled goat anti-rabbit at 1/2000 dilution
Predicted band size: 58 kDa
Observed band size: 165 kDa why is the actual band size different from the predicted? -
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PODXL antibody [EPR9518] (ab150358)
Immunohistochemical analysis of paraffin embedded human kidney tissue labeling PODXL with unpurified ab150358 antibody at a dilution of 1/100. Heat mediated antigen retrieval was performed before commencing with IHC staining protocol.
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Intracellular Flow Cytometry analysis of HeLa (human cervix adenocarcinoma) cells labeling PODXL with purified ab150358 at 1/250 dilution (red). The secondary antibody was Goat anti rabbit IgG (Alexa Fluor® 488) at 1/2000 dilution. A Rabbit monoclonal IgG (Black) was used as the isotype control and cells without incubation with primary antibody and secondary antibody (Blue) were used as unlabeled control.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PODXL antibody [EPR9518] (ab150358)This image is courtesy of an anonymous abreview.
Unpurified ab150358 staining PODXL in human kidney tissue sections by Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections). Tissue samples were fixed with formaldehyde, cut into 20 micron slices, permeabilized with 0.05% tween-20 and blocked for 60 minutes at 25°C. Antigen retrieval was by heat mediation. The sample was incubated with primary antibody at a dilution of 1/250 at 25°C for 1 hour. An Alexa Fluor® 488-conjugated donkey anti-rabbit polyclonal (1/1000) was used as the secondary antibody, at a dilution of 1/1200.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PODXL antibody [EPR9518] (ab150358)
Immunohistochemical analysis of paraffin embedded human hepatocellular carcinoma vessels using unpurified ab150358 showing positive staining. Heat mediated antigen retrieval was performed before commencing with IHC staining protocol.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PODXL antibody [EPR9518] (ab150358)
Immunohistochemical analysis of paraffin embedded human breast adenocarcinoma tissue using unpurified ab150358 showing positive staining. Heat mediated antigen retrieval was performed before commencing with IHC staining protocol.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PODXL antibody [EPR9518] (ab150358)
Immunohistochemical analysis of paraffin embedded human endometrial carcinoma tissue using unpurified ab150358 showing positive staining. Heat mediated antigen retrieval was performed before commencing with IHC staining protocol.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PODXL antibody [EPR9518] (ab150358)
Immunohistochemical analysis of paraffin embedded human skeletal muscle tissue using unpurified ab150358 showing negative staining. Heat mediated antigen retrieval was performed before commencing with IHC staining protocol.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PODXL antibody [EPR9518] (ab150358)
Immunohistochemical analysis of paraffin embedded normal human kidney tissue using unpurified ab150358 showing positive staining. Heat mediated antigen retrieval was performed before commencing with IHC staining protocol.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PODXL antibody [EPR9518] (ab150358)
Immunohistochemical analysis of paraffin embedded human glioma tissue using unpurified ab150358 showing positive staining. Heat mediated antigen retrieval was performed before commencing with IHC staining protocol.
Protocols
Datasheets and documents
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SDS download
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Datasheet download
References (9)
ab150358 has been referenced in 9 publications.
- Dhondt B et al. Unravelling the proteomic landscape of extracellular vesicles in prostate cancer by density-based fractionation of urine. J Extracell Vesicles 9:1736935 (2020). PubMed: 32284825
- Zhai S et al. IL-17 aggravates renal injury by promoting podocyte injury in children with primary nephrotic syndrome. Exp Ther Med 20:409-417 (2020). PubMed: 32537005
- Ito S et al. Identification of Cell-Surface Proteins Endocytosed by Human Brain Microvascular Endothelial Cells In Vitro. Pharmaceutics 12:N/A (2020). PubMed: 32585920
- Zhang D et al. In vitro induction and in vivo engraftment of kidney organoids derived from human pluripotent stem cells. Exp Ther Med 20:1307-1314 (2020). PubMed: 32742364
- Dhondt B et al. Preparation of Multi-omics Grade Extracellular Vesicles by Density-Based Fractionation of Urine. STAR Protoc 1:100073 (2020). PubMed: 33111109
- Wong BS et al. A Direct Podocalyxin-Dynamin-2 Interaction Regulates Cytoskeletal Dynamics to Promote Migration and Metastasis in Pancreatic Cancer Cells. Cancer Res 79:2878-2891 (2019). PubMed: 30975647
- Zhi Q et al. Podocalyxin-like protein promotes gastric cancer progression through interacting with RUN and FYVE domain containing 1 protein. Cancer Sci 110:118-134 (2019). PubMed: 30407695
- Zhang LH et al. Early podocyte injury and elevated levels of urinary podocyte-derived extracellular vesicles in swine with metabolic syndrome: role of podocyte mitochondria. Am J Physiol Renal Physiol 317:F12-F22 (2019). PubMed: 31042059
- Wang P et al. Dissecting the Global Dynamic Molecular Profiles of Human Fetal Kidney Development by Single-Cell RNA Sequencing. Cell Rep 24:3554-3567.e3 (2018). PubMed: 30257215