Recombinant Anti-IL-1 beta antibody [EPR23851-127] (ab254360)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR23851-127] to IL-1 beta
- Suitable for: Flow Cyt, ICC/IF, WB
- Knockout validated
- Reacts with: Mouse, Rat, Human
Related conjugates and formulations
Overview
-
Product name
Anti-IL-1 beta antibody [EPR23851-127]
See all IL-1 beta primary antibodies -
Description
Rabbit monoclonal [EPR23851-127] to IL-1 beta -
Host species
Rabbit -
Specificity
IL-1 beta is not present under homeostatic conditions, and it is induced and secreted only upon inflammatory signals. Stimulation may be required for the detection of IL-1β, as it is not constitutively expressed, no matter in precursor or mature form. For example, when using ab254360, positive signal can be detected in RAW264.7 with 100 ng/ml LPS treatment for 7h and additional 300 ng/ml Brefeldin A treatment in the last 3h. IL-1β is a secreted protein, it is recommended to treat cells with anti-secretion reagents (eg. Brefeldin A) that inhibit secretion.
-
Tested applications
Suitable for: Flow Cyt, ICC/IF, WBmore details
Unsuitable for: IHC-P or IP -
Species reactivity
Reacts with: Mouse, Rat, Human -
Immunogen
Recombinant fragment. This information is proprietary to Abcam and/or its suppliers.
-
Positive control
- WB: RAW 264.7 treated with 100 ng/ml LPS for 7 h and 300 ng/ml Brefeldin A for the last 3h, THP-1 treated with 80 nM TPA overnight and then 100 ng/ml LPS for 6 h and 300 ng/ml Brefeldin A for the last 3 h, NR8383 treated with 100 ng/ml LPS for 7 h and 300 ng/ml Brefeldin A for the last 3 h lysates. ICC/IF: RAW 264.7 cells; THP-1 cells. Flow Cyt: THP-1 and RAW 264.7, NR8383 cells.
-
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
-
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
pH: 7.2
Preservative: 0.01% Sodium azide
Constituents: 59.94% PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA -
Concentration information loading...
-
Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
EPR23851-127 -
Isotype
IgG -
Research areas
Associated products
-
Alternative Versions
-
Compatible Secondaries
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab254360 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 |
---|---|---|
Flow Cyt |
1/50.
|
|
ICC/IF |
1/50.
|
|
WB |
1/1000. Predicted molecular weight: 30 kDa.
|
Notes |
---|
Flow Cyt
1/50. |
ICC/IF
1/50. |
WB
1/1000. Predicted molecular weight: 30 kDa. |
Target
-
Function
Potent proinflammatory cytokine. Initially discovered as the major endogenous pyrogen, induces prostaglandin synthesis, neutrophil influx and activation, T-cell activation and cytokine production, B-cell activation and antibody production, and fibroblast proliferation and collagen production. Promotes Th17 differentiation of T-cells. -
Tissue specificity
Expressed in activated monocytes/macrophages (at protein level). -
Sequence similarities
Belongs to the IL-1 family. -
Post-translational
modificationsActivation of the IL1B precursor involves a CASP1-catalyzed proteolytic cleavage. Processing and secretion are temporarily associated. -
Cellular localization
Cytoplasm, cytosol. Lysosome. Secreted, exosome. Cytoplasmic vesicle, autophagosome. Secreted. The precursor is cytosolic. In response to inflammasome-activating signals, such as ATP for NLRP3 inflammasome or bacterial flagellin for NLRC4 inflammasome, cleaved and secreted. IL1B lacks any known signal sequence and the pathway(s) of its secretion is(are) not yet fully understood (PubMed:24201029). On the basis of experimental results, several unconventional secretion mechanisms have been proposed. 1. Secretion via secretory lysosomes: a fraction of CASP1 and IL1B precursor may be incorporated, by a yet undefined mechanism, into secretory lysosomes that undergo Ca(2+)-dependent exocytosis with release of mature IL1B (PubMed:15192144). 2. Secretory autophagy: IL1B-containing autophagosomes may fuse with endosomes or multivesicular bodies (MVBs) and then merge with the plasma membrane releasing soluble IL1B or IL1B-containing exosomes (PubMed:24201029). However, autophagy impacts IL1B production at several levels and its role in secretion is still controversial. 3. Secretion via exosomes: ATP-activation of P2RX7 leads to the formation of MVBs containing exosomes with entrapped IL1B, CASP1 and other inflammasome components. These MVBs undergo exocytosis with the release of exosomes. The release of soluble IL1B occurs after the lysis of exosome membranes (By similarity). 4. Secretion by microvesicle shedding: activation of the ATP receptor P2RX7 may induce an immediate shedding of membrane-derived microvesicles containing IL1B and possibly inflammasome components. The cytokine is then released in the extracellular compartment after microvesicle lysis (PubMed:11728343). 5. Release by translocation through permeabilized plasma membrane. This may occur in cells undergoing pyroptosis due to sustained activation of the inflammasome (By similarity). These mechanisms may not be not mutually exclusive. - Information by UniProt
-
Database links
- Entrez Gene: 3553 Human
- Entrez Gene: 16176 Mouse
- Entrez Gene: 24494 Rat
- Omim: 147720 Human
- SwissProt: P01584 Human
- SwissProt: P10749 Mouse
- SwissProt: Q63264 Rat
- Unigene: 126256 Human
see all -
Alternative names
- Catabolin antibody
- H1 antibody
- IFN beta inducing factor antibody
see all
Images
-
ab254360 staining IL-1 beta in wild-type THP-1 cells and IL1B knockout THP-1 cells (ab273762) +/- TPA (80nM overnight) and LPS (100ng/ml, 6h). The cells were fixed with 4% paraformaldehyde (10 min) then permeabilized with 0.1% 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 with ab254360 at 0.4μg/ml concentration and ab7291 (Mouse monoclonal to alpha Tubulin) at 1/1000 dilution overnight at 4°C followed by a further incubation at room temperature for 1h with a goat secondary antibody to rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green) and a goat secondary antibody to mouse IgG (Alexa Fluor® 594) (ab150120) at 2 μg/ml (shown in red). Nuclear DNA was labelled in blue with DAPI.
Image was taken with a confocal microscope (Leica-Microsystems TCS SP8). -
All lanes : Anti-IL-1 beta antibody [EPR23851-127] (ab254360) at 1/1000 dilution
Lane 1 : Untreated RAW 264.7 (mouse abelson murine leukemia virus-induced tumor macrophage) whole cell lysate
Lane 2 : RAW 264.7 treated with 100 ng/ml LPS for 7 hours and 300 ng/ml Brefeldin A (ab193369) for the last 3 hours, whole cell lysate
Lane 3 : Untreated THP-1 (human monocytic leukemia monocyte) whole cell lysate
Lane 4 : THP-1 treated with 80 nM TPA (ab120297) overnight and then 100 ng/ml LPS for 6 hours and 300 ng/ml Brefeldin A (ab193369) for the last 3 hours, whole cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/50000 dilution (Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated)
Predicted band size: 30 kDa
Observed band size: 31/28/17.5 kDa why is the actual band size different from the predicted?Blocking and diluting buffer and concentration: 5% NFDM/TBST
Expression of IL-1 beta is induced by LPS treatment. 31-kDa precursor IL-1 beta, 28- and 17.5-kDa proteolytically cleaved IL-1 beta are observed. The expesssion pattern and molecular weight observed is consistent with what has been described in the literature (PMID: 8446594, 19559631).
Exposure time: 3 minutes
-
All lanes : Anti-IL-1 beta antibody [EPR23851-127] (ab254360) at 1/1000 dilution
Lane 1 : Untreated NR8383 (rat lung macrophage (alveolar)) whole cell lysate
Lane 2 : NR8383 treated with 100 ng/ml LPS for 7 hours and 300 ng/ml Brefeldin A (ab193369) for the last 3 hours, whole cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/50000 dilution (Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated)
Predicted band size: 30 kDa
Observed band size: 31/28/17.5 kDa why is the actual band size different from the predicted?Blocking and diluting buffer and concentration: 3% NFDM/TBST
Nitrocellulose membrane was used in this blot.
Exposure time: 70 seconds
-
Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized RAW 264.7 cells labelling IL-1 beta with ab254360 at 1/50 (10.28 µg/ml) dilution, followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) antibody at 1/1000 dilution (Green) Confocal image showing increased cytoplasmic and nuclear staining in RAW 264.7 cells treated with Lipopolysaccharide (100 ng/ml) for 4 h then together with Brefeldin A (1 µg/ml) for another 3 h is observed.
ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 dilution (Red). The Nuclear counterstain was DAPI (Blue).
Secondary antibody only control: Secondary antibody is ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/1000 dilution.
-
Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized THP-1(Human monocytic leukemia monocyte) treated with 80nM TPA for 16 hours, then 100ng/ml LPS for 3 hours and 300ng/ml BFA for another 3 hours (Right)/ Untreated control (Left) cells labelling IL-1 beta with ab254360 at 1/500 dilution (0.1ug). A Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) at 1/2000 dilution was used as the secondary antibody.
-
Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized RAW 264.7 (Mouse Abelson murine leukemia virus-induced tumor macrophage) treated with 100ng/ml LPS for 4 hours and 1ug/ml BFA for another 3 hours (Right)/ Untreated control (Left) cells labelling IL-1 beta with ab254360 at 1/500 dilution (0.1ug). A Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) at 1/2000 dilution was used as the secondary antibody.
-
Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized NR8383 (rat lung macrophage (alveolar)) treated with 100ng/ml LPS for 4 hours and 1ug/ml BFA for another 3 hours (Right)/ Untreated control (Left) cells labelling IL-1 beta with ab254360 at 1/50 dilution (1ug)/ Right and Left. A Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) at 1/2000 dilution was used as the secondary antibody.
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
-
SDS download
-
Datasheet download
Certificate of Compliance
References (2)
ab254360 has been referenced in 2 publications.
- Li L et al. Tetrahydrocurcumin protects against sepsis-induced acute kidney injury via the SIRT1 pathway. Ren Fail 43:1028-1040 (2021). PubMed: 34187277
- Jiang Z et al. Anti-inflammatory effects of paeoniflorin caused by regulation of the hif1a/miR-210/caspase1/GSDMD signaling pathway in astrocytes: a novel strategy for hypoxia-induced brain injury in rats. Immunopharmacol Immunotoxicol 43:410-418 (2021). PubMed: 34114917