Recombinant Anti-P4HB antibody [EPR9499] (ab137110)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR9499] to P4HB
- Suitable for: Flow Cyt (Intra), WB, IHC-P, ICC/IF
- Knockout validated
- Reacts with: Mouse, Rat, Human
Related conjugates and formulations
Overview
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Product name
Anti-P4HB antibody [EPR9499]
See all P4HB primary antibodies -
Description
Rabbit monoclonal [EPR9499] to P4HB -
Host species
Rabbit -
Tested applications
Suitable for: Flow Cyt (Intra), WB, IHC-P, ICC/IFmore details -
Species reactivity
Reacts with: Mouse, Rat, Human -
Immunogen
Synthetic peptide within Human P4HB aa 450 to the C-terminus (C terminal). The exact sequence is proprietary.
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Positive control
- WB: HeLa, HepG2, 293T, Raw264.7, PC-12 and A431 cell lysates. ICC/IF: HeLa, A-431, MCF7, A549 and U2OS cells IHC-P: Human breast carcinoma, human brain, human kidney, mouse liver and rat liver tissues. Flow Cyt (intra): HepG2 and 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.
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: 40% Glycerol, 0.05% BSA, 59% PBS -
Concentration information loading...
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Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
EPR9499 -
Isotype
IgG -
Research areas
Associated products
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Alternative Versions
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Compatible Secondaries
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Conjugation kits
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Isotype control
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Positive Controls
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab137110 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) |
1/100 - 1/1000.
ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody. |
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WB | (3) |
1/1000 - 1/10000. Predicted molecular weight: 57 kDa.
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IHC-P |
1/1000. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
See IHC antigen retrieval protocols. For unpurified use at 1/100 - 1/250. |
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ICC/IF | (3) |
1/150 - 1/500.
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Notes |
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Flow Cyt (Intra)
1/100 - 1/1000. ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody. |
WB
1/1000 - 1/10000. Predicted molecular weight: 57 kDa. |
IHC-P
1/1000. Perform heat mediated antigen retrieval before commencing with IHC staining protocol. See IHC antigen retrieval protocols. For unpurified use at 1/100 - 1/250. |
ICC/IF
1/150 - 1/500. |
Target
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Function
This multifunctional protein catalyzes the formation, breakage and rearrangement of disulfide bonds. At the cell surface, seems to act as a reductase that cleaves disulfide bonds of proteins attached to the cell. May therefore cause structural modifications of exofacial proteins. Inside the cell, seems to form/rearrange disulfide bonds of nascent proteins. At high concentrations, functions as a chaperone that inhibits aggregation of misfolded proteins. At low concentrations, facilitates aggregation (anti-chaperone activity). May be involved with other chaperones in the structural modification of the TG precursor in hormone biogenesis. Also acts a structural subunit of various enzymes such as prolyl 4-hydroxylase and microsomal triacylglycerol transfer protein MTTP. -
Sequence similarities
Belongs to the protein disulfide isomerase family.
Contains 2 thioredoxin domains. -
Cellular localization
Endoplasmic reticulum lumen. Melanosome. Cell membrane. Highly abundant. In some cell types, seems to be also secreted or associated with the plasma membrane, where it undergoes constant shedding and replacement from intracellular sources (Probable). Localizes near CD4-enriched regions on lymphoid cell surfaces. Identified by mass spectrometry in melanosome fractions from stage I to stage IV. - Information by UniProt
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Database links
- Entrez Gene: 5034 Human
- Entrez Gene: 18453 Mouse
- Entrez Gene: 25506 Rat
- Omim: 176790 Human
- SwissProt: P07237 Human
- SwissProt: P09103 Mouse
- SwissProt: P04785 Rat
- Unigene: 464336 Human
see all -
Alternative names
- Cellular thyroid hormone binding protein antibody
- Cellular thyroid hormone-binding protein antibody
- Collagen prolyl 4 hydroxylase beta antibody
see all
Images
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All lanes : Anti-P4HB antibody [EPR9499] (ab137110) at 1/1000 dilution
Lane 1 : HeLa cell lysate
Lane 2 : HepG2 cell lysate
Lane 3 : Wild-type A431 cell lysate
Lane 4 : P4HB knockout A431 cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 57 kDa
Observed band size: 60 kDa why is the actual band size different from the predicted?False colour image of Western blot: Anti-P4HB antibody [EPR9499] staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab137110 was shown to bind specifically to P4HB. A band was observed at 60 kDa in wild-type HeLa cell lysates with no signal observed at this size in P4HB knockout cell line ab261887 (knockout cell lysate ab261696). To generate this image, wild-type and P4HB knockout HeLa cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3% milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4°C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) at 1/20000 dilution.
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ab137110 staining P4HB in wild-type A431 cells (top panel) and P4HB knockout A431 cells (bottom panel). The cells were fixed with 100% methanol (5 min), permeabilized with 0.1% PBS-Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1h. The cells were then incubated overnight at 4°C with ab137110 at 1µg/ml and ab7291, Mouse monoclonal [DM1A] to alpha Tubulin - Loading Control. Cells were then incubated with ab150081, Goat polyclonal Secondary Antibody to Rabbit IgG - H&L (Alexa Fluor® 488), pre-adsorbed at 1/1000 dilution (shown in green) and ab150120, Goat polyclonal Secondary Antibody to Mouse IgG - H&L (Alexa Fluor® 594), pre-adsorbed at 1/1000 dilution (shown in pseudocolour red). Nuclear DNA was labelled with DAPI (shown in blue).
Also suitable in cells fixed with 4% paraformaldehyde (10 min).
Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a maximum intensity projection of confocal sections is shown.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-P4HB antibody [EPR9499] (ab137110)
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human kidney carcinoma tissue labelling P4HB with purified ab137110 at a dilution of 1/1000. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. ab97051, a HRP-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.
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ab137110 staining P4HB in U2OS cells. The cells were fixed with 100% methanol (5 min), permeabilized with 0.1% PBS-Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1h. The cells were then incubated overnight at 4°C with ab137110 at 0.2µg/ml and ab7291, Mouse monoclonal [DM1A] to alpha Tubulin - Loading Control. Cells were then incubated with ab150081, Goat polyclonal Secondary Antibody to Rabbit IgG - H&L (Alexa Fluor® 488), pre-adsorbed at 1/1000 dilution (shown in green) and ab150120, Goat polyclonal Secondary Antibody to Mouse IgG - H&L (Alexa Fluor® 594), pre-adsorbed at 1/1000 dilution (shown in pseudocolour red). Nuclear DNA was labelled with DAPI (shown in blue).
Also suitable in cells fixed with 4% paraformaldehyde (10 min).
Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a maximum intensity projection of confocal sections is shown.
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ab137110 staining P4HB in A549 cells. The cells were fixed with 100% methanol (5 min), permeabilized with 0.1% PBS-Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1h. The cells were then incubated overnight at 4°C with ab137110 at 0.2µg/ml and ab7291, Mouse monoclonal [DM1A] to alpha Tubulin - Loading Control. Cells were then incubated with ab150081, Goat polyclonal Secondary Antibody to Rabbit IgG - H&L (Alexa Fluor® 488), pre-adsorbed at 1/1000 dilution (shown in green) and ab150120, Goat polyclonal Secondary Antibody to Mouse IgG - H&L (Alexa Fluor® 594), pre-adsorbed at 1/1000 dilution (shown in pseudocolour red). Nuclear DNA was labelled with DAPI (shown in blue).
Also suitable in cells fixed with 4% paraformaldehyde (10 min).
Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a maximum intensity projection of confocal sections is shown.
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ab137110 staining P4HB in HeLa cells. The cells were fixed with 100% methanol (5 min), permeabilized with 0.1% PBS-Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1h. The cells were then incubated overnight at 4°C with ab137110 at 0.2µg/ml and ab7291, Mouse monoclonal [DM1A] to alpha Tubulin - Loading Control. Cells were then incubated with ab150081, Goat polyclonal Secondary Antibody to Rabbit IgG - H&L (Alexa Fluor® 488), pre-adsorbed at 1/1000 dilution (shown in green) and ab150120, Goat polyclonal Secondary Antibody to Mouse IgG - H&L (Alexa Fluor® 594), pre-adsorbed at 1/1000 dilution (shown in pseudocolour red). Nuclear DNA was labelled with DAPI (shown in blue).
Also suitable in cells fixed with 4% paraformaldehyde (10 min).
Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a maximum intensity projection of confocal sections is shown.
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ab137110 staining P4HB in MCF7 cells. The cells were fixed with 100% methanol (5 min), permeabilized with 0.1% PBS-Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1h. The cells were then incubated overnight at 4°C with ab137110 at 1µg/ml and ab7291, Mouse monoclonal [DM1A] to alpha Tubulin - Loading Control. Cells were then incubated with ab150081, Goat polyclonal Secondary Antibody to Rabbit IgG - H&L (Alexa Fluor® 488), pre-adsorbed at 1/1000 dilution (shown in green) and ab150120, Goat polyclonal Secondary Antibody to Mouse IgG - H&L (Alexa Fluor® 594), pre-adsorbed at 1/1000 dilution (shown in pseudocolour red). Nuclear DNA was labelled with DAPI (shown in blue).
Also suitable in cells fixed with 4% paraformaldehyde (10 min).
Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a maximum intensity projection of confocal sections is shown.
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All lanes : Anti-P4HB antibody [EPR9499] (ab137110) at 1/10000 dilution (purified)
Lane 1 : HepG2 whole cell lysate
Lane 2 : HeLa whole cell lysate
Lane 3 : HEK293 whole cell lysate
Lysates/proteins at 10 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Predicted band size: 57 kDa
Observed band size: 57 kDa
Blocking and dilution buffer: 5% NFDM/TBST -
All lanes : Anti-P4HB antibody [EPR9499] (ab137110) at 1/10000 dilution (purified)
Lane 1 : Raw264.7 whole cell lysate
Lane 2 : PC-12 whole cell lysate
Lysates/proteins at 10 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Predicted band size: 57 kDa
Observed band size: 57 kDa
Blocking and dilution buffer: 5% NFDM/TBST -
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-P4HB antibody [EPR9499] (ab137110)
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of mouse liver tissue labelling P4HB with purified ab137110 at a dilution of 1/1000. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. ab97051, a HRP-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-P4HB antibody [EPR9499] (ab137110)
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of rat liver tissue labelling P4HB with purified ab137110 at a dilution of 1/1000. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. ab97051, a HRP-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.
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Intracellular Flow Cytometry analysis ofHeLa cells labelling P4HB with purified ab137110 at a dilution of 1/300 (red). Cells were fixed with 4% paraformaldehyde. An Alexa Fluor®488-conjugated goat anti-rabbit IgG (1/500) was used as the secondary antibody. Black - Isotype control, rabbit monoclonal IgG. Blue - Unlabelled control, cells without incubation with primary and secondary antibodies.
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All lanes : Anti-P4HB antibody [EPR9499] (ab137110) at 1/1000 dilution (unpurified)
Lane 1 : HepG2 cell lysate
Lane 2 : HeLa cell lysate
Lane 3 : 293T cell lysate
Lane 4 : A431 cell lysate
Lysates/proteins at 10 µg per lane.
Secondary
All lanes : HRP-conjugated goat anti-rabbit IgG at 1/2000 dilution
Developed using the ECL technique.
Predicted band size: 57 kDa -
All lanes : Anti-P4HB antibody [EPR9499] (ab137110) at 1/2000 dilution (unpurified)
Lane 1 : Mouse brain tissue lysate
Lane 2 : Mouse heart tissue lysate
Lane 3 : Mouse kidney tissue lysate
Lane 4 : Mouse spleen tissue lysate
Predicted band size: 57 kDa -
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-P4HB antibody [EPR9499] (ab137110)
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human breast carcinoma tissue labelling P4HB with unpurified ab137110 at a dilution of 1/100.
Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-P4HB antibody [EPR9499] (ab137110)
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human brain tissue labelling P4HB with unpurified ab137110 at a dilution of 1/100.
Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
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Overlay histogram showing HepG2 cells stained with unpurified ab137110 (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 (unpurified ab137110, 1/1000 dilution) for 30 min at 22°C. The secondary antibody used was a goat anti-rabbit Alexa Fluor® 488 (IgG H&L) (ab150077) at 1/2000 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (0.1µg/1x106 cells) used under the same conditions. Unlabelled sample (blue line) was also used as a control. Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter. This antibody gave a positive signal in HepG2 cells fixed with 4% paraformaldehyde (10 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.
Protocols
Datasheets and documents
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SDS download
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Datasheet download
References (14)
ab137110 has been referenced in 14 publications.
- Zhong T et al. Key genes associated with prognosis and metastasis of clear cell renal cell carcinoma. PeerJ 10:e12493 (2022). PubMed: 35036081
- Wu Y et al. P4HB: A novel diagnostic and prognostic biomarker for bladder carcinoma. Oncol Lett 21:95 (2021). PubMed: 33376528
- Singhmar P et al. The fibroblast-derived protein PI16 controls neuropathic pain. Proc Natl Acad Sci U S A 117:5463-5471 (2020). PubMed: 32079726
- Guo H et al. miR-21 is upregulated, promoting fibrosis and blocking G2/M in irradiated rat cardiac fibroblasts. PeerJ 8:e10502 (2020). PubMed: 33354435
- Dhall S et al. A Flowable Placental Formulation Prevents Bleomycin-Induced Dermal Fibrosis in Aged Mice. Int J Mol Sci 21:N/A (2020). PubMed: 32545915