Recombinant Anti-IL-8 antibody [EPR26511-74] (ab289967)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR26511-74] to IL-8
- Suitable for: ICC/IF, Flow Cyt (Intra), IP, WB
- Knockout validated
- Reacts with: Human
Related conjugates and formulations
Overview
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Product name
Anti-IL-8 antibody [EPR26511-74]
See all IL-8 primary antibodies -
Description
Rabbit monoclonal [EPR26511-74] to IL-8 -
Host species
Rabbit -
Tested applications
Suitable for: ICC/IF, Flow Cyt (Intra), IP, WBmore details
Unsuitable for: IHC-P -
Species reactivity
Reacts with: Human -
Immunogen
Recombinant fragment. This information is proprietary to Abcam and/or its suppliers.
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Positive control
- WB: Wild-type treated PC-3, treated U-937, Untreated U-87 MG, U-87 MG treated with 1µM Thapsigargin for 24h. ICC/IF: Treated U-937 cells. Flow Cyt (intra): Treated U-937 cells, treated Human peripheral blood mononuclear cell (PBMC). IP: treated U-937 whole cell lysate
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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.
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 long term. Avoid freeze / thaw cycle. -
Storage buffer
pH: 7.20
Preservative: 0.01% Sodium azide
Constituents: 59% PBS, 0.05% BSA, 40% Glycerol (glycerin, glycerine) -
Concentration information loading...
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Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
EPR26511-74 -
Isotype
IgG -
Research areas
Associated products
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Alternative Versions
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Isotype control
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Related Products
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab289967 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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ICC/IF |
1/100.
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Flow Cyt (Intra) |
1/500.
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IP |
1/30.
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WB |
1/1000. Predicted molecular weight: 11 kDa.
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Notes |
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ICC/IF
1/100. |
Flow Cyt (Intra)
1/500. |
IP
1/30. |
WB
1/1000. Predicted molecular weight: 11 kDa. |
Target
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Function
IL-8 is a chemotactic factor that attracts neutrophils, basophils, and T-cells, but not monocytes. It is also involved in neutrophil activation. It is released from several cell types in response to an inflammatory stimulus. IL-8(6-77) has a 5-10-fold higher activity on neutrophil activation, IL-8(5-77) has increased activity on neutrophil activation and IL-8(7-77) has a higher affinity to receptors CXCR1 and CXCR2 as compared to IL-8(1-77), respectively. -
Sequence similarities
Belongs to the intercrine alpha (chemokine CxC) family. -
Post-translational
modificationsSeveral N-terminal processed forms are produced by proteolytic cleavage after secretion from at least peripheral blood monocytes, leukcocytes and endothelial cells. In general, IL-8(1-77) is referred to as interleukin-8. IL-8(6-77) is the most promiment form. -
Cellular localization
Secreted. - Information by UniProt
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Database links
- Entrez Gene: 3576 Human
- Omim: 146930 Human
- SwissProt: P10145 Human
- Unigene: 624 Human
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Alternative names
- (Ala-IL-8)77 antibody
- (Ser-IL-8)72 antibody
- 9E3 antibody
see all
Images
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All lanes : Anti-IL-8 antibody [EPR26511-74] (ab289967) at 1/1000 dilution
Lane 1 : Wild-type PC-3 (human prostate adenocarcinoma epithelial cell) whole cell lysate
Lane 2 : Wild-type PC-3 treated with 2µg/ml LPS for 5h, then treated with 5µg/ml Brefeldin A for 5h, whole cell lysate
Lane 3 : CXCL8 knockout PC-3 whole cell lysate
Lane 4 : CXCL8 knockout PC-3 treated with 2µg/ml LPS for 5h, then treated with 5µg/ml Brefeldin A for 5h, whole cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) at 1/10000 dilution
Predicted band size: 11 kDa
Observed band size: 11 kDaBlocking buffer and concentration was Intercept® (TBS) Blocking Buffer diluted with an equal volume of 0.1% TBS.
Diluting buffer was Intercept® (TBS) Blocking Buffer diluted with an equal volume of 0.1% TBST.
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
False colour image of Western blot: Anti-IL-8 antibody [EPR26511-74] (ab289967) 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, ab289967 was shown to bind specifically to IL-8. A band was observed at 11 kDa in wild-type PC-3 cell lysates with no signal observed at this size in CXCL8 knockout cell lysates. To generate this image, wild-type and CXCL8 knockout PC-3 cell lysates were analyzed. First, samples were run on an SDS-PAGE gel then transferred onto an immobilon-FL PVDF membrane. Membranes were blocked in Intercept® (TBS) Blocking Buffer diluted with an equal volume of 0.1% TBS 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 1/10000 dilution. -
All lanes : Anti-IL-8 antibody [EPR26511-74] (ab289967) at 1/1000 dilution
Lane 1 : Untreated U-937 (human histiocytic lymphoma monocyte) whole cell lysate
Lane 2 : U-937 treated with TPA (100ng/mL) for 24 h, then treated with LPS (5 µg/mL) for 7 h with Brefeldin A (300 ng/mL) for the last 3 h, whole cell lysate
Lane 3 : Untreated U-87 MG (human glioblastoma-astrocytoma epithelial cell) whole cell lysate
Lane 4 : U-87 MG treated with 1µM Thapsigargin for 24h, whole cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution
Predicted band size: 11 kDa
Observed band size: 11 kDa
Exposure time: 70 secondsBlocking and diluting buffer and concentration was 5% NFDM/TBST.
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Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized U-937 cells labelling IL-8 with ab289967 at 1/100 (5.87 µg/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 (2 µg/mL) dilution (Green). Confocal image showing cytoplasmic staining is observed in U-937 cells treated with TPA (100 ng/mL) for 24 h, then LPS (5 µg/mL) for 7 h with Brefeldin A (300 ng/mL) for the last 3 h. ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 (2.5µg/ml) dilution (Red). The Nuclear counterstain was DAPI (Blue).
Secondary antibody only control: Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 (2 µg/mL) dilution.
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Intracellular flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized U-937 (Human histiocytic lymphoma monocyte) treated with 100ng/ml TPA for 24 hours, then 5µg/ml LPS for 4 hours, and add 300ng/ml BFA for another 3h (Red) / Untreated control (Green) cells labelling IL-8 with ab289967 at 1/500 dilution (0.1µg) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). A Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/2000 dilution was used as the secondary antibody.
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Intracellular flow cytometric analysis of 2% paraformaldehyde fixed, 0.1% saponin permeabilised human peripheral blood mononuclear cell (PBMC) treated with 1μg/ml Lipopolysaccharide (LPS) for 22 hours, then add 3uM Monensin for another 2h (Right). Untreated control (Left).
Primary antibody: ab289967, at 1/500 dilution.
Secondary antibody: Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/2000 dilution.
Scatter image shows specific IL-8 expression in LPS induced monocyte population.
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IL-8 was immunoprecipitated from U937 (human histiocytic lymphoma monocyte) treated with 100ng/ml TPA for 24h then treated with 5 µg/ml LPS for 4h, and add 300 ng/ml BFA for another 3h, whole cell lysate with ab289967 at 1/30 dilution (2 µg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab289967 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.
Lane 1: U937 (human histiocytic lymphoma monocyte) treated with 100 ng/ml TPA for 24h then treated with 5 µg/ml LPS for 4h, and add 300 ng/ml BFA for another 3h, whole cell lysate 10 µg
Lane 2: ab289967 IP in U937 treated with 100 ng/ml TPA for 24h then treated with 5 µg/ml LPS for 4h, and add 300 ng/ml BFA for another 3h, whole cell lysate
Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab289967 in U937 treated with 100 ng/ml TPA for 24h then treated with 5 µg/ml LPS for 4h, and add 300 ng/ml BFA for another 3h, whole cell lysate
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 50 seconds
Protocols
To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.
Datasheets and documents
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SDS download
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Datasheet download
Certificate of Compliance
References (1)
ab289967 has been referenced in 1 publication.
- Piao H et al. A positive feedback loop between gastric cancer cells and tumor-associated macrophage induces malignancy progression. J Exp Clin Cancer Res 41:174 (2022). PubMed: 35562774