Overview

  • Product name
  • Description
    Rabbit polyclonal to IL-1 beta
  • Host species
    Rabbit
  • Tested applications
    Suitable for: WB, IP, ELISA, IHC-P, RIA, IHC-Fr, Functional Studies, Flow Cyt, Neutralising, ICC/IF, IHC-FoFrmore details
  • Species reactivity
    Reacts with: Human
    Predicted to work with: Non human primates
  • Immunogen

    Recombinant 153 aa human IL-1beta produced in E.coli. MW of recombinant IL-1beta 17kDa, with the N-terminal amino acid at position alanine 117. This is the cleavage site generated by the IL-1beta converting enzyme (ICE, capase-1).

  • General notes
    Immunohistochemistry 1/100 to 1/200 (paraffin or cryofixation can be used; 1/100 for staining intracellular IL-1beta). ELISA 1/200 to 1/1000 (this antibody is best used as the second antibody with a monoclonal as a capture antibody). Radio immunoassay 1/8000 Neutralization of IL-1beta activity in bioassays 1/100 ( >4 hours incubation, normal rabbit IgG as negative control) FACS analysis - caution, F(c) domain of rabbit IgG may interact with cells non-specifically.

Properties

Applications

Our Abpromise guarantee covers the use of ab2105 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
WB Use at an assay dependent concentration.

1/1000 - 1/2000 (antigen - supernatants or lysates of 2 x 106 endotoxin-stimulated human PBMC. Denatured 31kDa precursor IL-1beta will be recognized, but often migrates as a 35 kDa band).

IP Use at an assay dependent concentration. 1/400 - 1/800 (pre-clearing with a non-specific rabbit IgG is helpful to reduce background).
ELISA 1/500 - 1/2000.
IHC-P 1/100 - 1/200.
RIA 1/8000.
IHC-Fr 1/100 - 1/200.
Functional Studies Use at an assay dependent concentration.
Flow Cyt Use at an assay dependent concentration.

F(c) domain of rabbit IgG may interact with cells non-specifically!

ab171870 - Rabbit polyclonal IgG, is suitable for use as an isotype control with this antibody.

Neutralising 1/100. Neutralization of IL-1beta activity in bioassays 1/100 ( >4 hours incubation, normal rabbit IgG as negative control). ab2105 does not neutralize the biological activity of murine, rat or rabbit IL1 beta.
ICC/IF Use at an assay dependent concentration.
IHC-FoFr Use at an assay dependent concentration. PubMed: 28726778

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
    modifications
    Activation 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
  • Alternative names
    • Catabolin antibody
    • H1 antibody
    • IL 1 antibody
    • IL 1 beta antibody
    • IL-1 beta antibody
    • IL1 BETA antibody
    • IL1B antibody
    • IL1B_HUMAN antibody
    • IL1F2 antibody
    • Interleukin 1 beta antibody
    • Interleukin-1 beta antibody
    • OAF antibody
    • OTTHUMP00000162031 antibody
    • Preinterleukin 1 beta antibody
    • Pro interleukin 1 beta antibody
    see all

Images

  • All lanes : Anti-IL-1 beta antibody (ab2105) at 1/1000 dilution

    Lane 1 : Human peripheral blood mononuclear cell lysate - PBMC's unstimulated
    Lane 2 : Human peripheral blood mononuclear cell lysate - monocytes unstimulated
    Lane 3 : Human peripheral blood mononuclear cell lysate - monocytes stimulated with LPS
    Lane 4 : Human peripheral blood mononuclear cell lysate - positive control (rh IL-1 beta protein)

    Lysates/proteins at 25 µg per lane.

    Secondary
    All lanes : HRP-conjugated Goat anti-rabbit IgG polyclonal at 1/2000 dilution

    Performed under reducing conditions.

    Observed band size: 35 kDa
    why is the actual band size different from the predicted?


    Exposure time: 10 minutes

    See Abreview

  • ab2105 at 1/20 staining human liver tissue sections by IHC-P. The cells were paraformaldehyde fixed and a heat mediated antigen retrieval step was performed. The tissue was incubated with the tissue overnight at room temperature. A biotinylated donkey anti-rabbit IgG was used as the secondary antibody and this was detected using streptavidin HRP.

    See Abreview

  • ab2105 staining IL-1 beta in human keratinocytes by Immunocytochemistry/ Immunofluorescence. Cells were fixed in methanol, permeabilized in 0,1% Saponin / PBS and then blocked using 4% BSA for 30 minutes at 25°C. Samples were then incubated with ab2105 at a 1/100 dilution for 2 hours at 4°C. An Alexa-fluor 488 conjugated goat anti-rabbit polyclonal was used as the secondary antibody at a 1/100 dilution.

    See Abreview

  • ab2105 staining IL-1 beta in Human monocytes by Flow Cytometry. Cells were fixed with paraformaldehyde and permeabilized with 0.1% saponin in PBS. The sample was incubated with the primary antibody (1/500 in 0.1% saponin + 1% FCS in PBS) for 45 minutes at 4°C. An Alexa Fluor® 488-conjugated Goat anti-rabbit IgG (1/100) was used as the secondary antibody.

    1 - unstimulated monocytes. 2 - monocytes stimulated with LPS.

    See Abreview

  • Lanes 2-5 : Anti-IL-1 beta antibody (ab2105) at 1/1000 dilution

    Lane 1 : Ladder (top to bottom: p35, p25, p20, p17, p11)
    Lane 2 : Human THP-1 leukemic monocyte cell lysate, no LPS at 20 µg
    Lane 3 : Human THP-1 leukemic monocyte supernatant, no LPS at 20 µg
    Lane 4 : Human THP-1 leukemic monocyte cell lysate, LPS stimulation at 20 µg
    Lane 5 : Human THP-1 leukemic monocyte supernatant, LPS stimulation at 20 µg

    Secondary
    Lanes 2-5 : IRDye® 800CW-conjugated Donkey anti-rabbit IgG monoclonal at 1/5000 dilution

    Performed under reducing conditions.

    Observed band size: 17,30 kDa why is the actual band size different from the predicted?


    Exposure time: 30 seconds


    Both mature and pro-IL-1beta bands, p17 and p31 were detected by ab2105 in supernatant from THP-1 cells stimulated with LPS. Only pro-IL-1beta band (p31) was detected in supernatant from THP-1 cells not stimulated with LPS.

    See Abreview

References

This product has been referenced in:
  • Wang S  et al. Oxidative stress, autophagy and pyroptosis in the neovascularization of oxygen-induced retinopathy in mice. Mol Med Rep 19:927-934 (2019). Read more (PubMed: 30569132) »
  • Wang X  et al. Effect of Gastrodin on Early Brain Injury and Neurological Outcome After Subarachnoid Hemorrhage in Rats. Neurosci Bull N/A:N/A (2019). Read more (PubMed: 30673960) »
See all 57 Publications for this product

Customer reviews and Q&As

31-38 of 38 Abreviews or Q&A

Application
Western blot
Sample
Human Cell lysate - nuclear (keratinocytes and tumoral cervical cell line)
Gel Running Conditions
Reduced Denaturing (15%)
Loading amount
50 µg
Specification
keratinocytes and tumoral cervical cell line
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Dec 04 2010

Application
Immunohistochemistry (Frozen sections)
Sample
Human Tissue sections (Intestine)
Permeabilization
No
Specification
Intestine
Blocking step
Serum as blocking agent for 30 minute(s) · Concentration: 10% · Temperature: 26°C
Fixative
Formaldehyde

Abcam user community

Verified customer

Submitted Aug 29 2007

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Human Tissue sections (liver)
Antigen retrieval step
Heat mediated
Specification
liver
Blocking step
No blocking step used for 10 minute(s) · Concentration: 1%
Fixative
Paraformaldehyde

Miss. Silke Vorwald

Verified customer

Submitted Mar 09 2007

Answer

Thank you for getting back to me. The standard advice is always to reduce your samples using mercaptoethanol, unless the data sheet specifies that the antibody recognises the non-reduced form of the protein, which ab2105 does not. If you are concerned you are going to use up a lot of antibody using higher dilutions, you could try the other optimisation suggestions first to see if they produce a good result. I hope that makes sense! Do get back to me if you have any more problems. With thanks

Read More

Answer

Thank you for your enquiry and taking the time to fill in the questionaire. I am sorry you have had trouble using this antibody for your western blotting. I have looked at the details of your procedure. There are a few points of the procedure that may need optimising. You are using the antibody at the highest recommended dilution. You could try titrating this down to 1:100 or even 1:50. Make sure you are storing your antbody correctly as stated on the data sheet by aliquoting it and storing at -20oC. You can then keep one working aliquot at 4oC for a short period of time. Keeping an antibody at 4oC for a long period and repeat freeze-thawing can mean the antibody is no longer useable. You could also optimise your blocking conditins. You have not stated the blocking incubation time or temperature, but you could try shortening the incubation time, incubating at 4oC and perhaps using 2% milk for a more gentle block. Your secondary antibody may also require titrating to work with this particular primary antibody. I hope this helps. Please get back to me if you require any more advice or if you have any more problems with this procedure. Good luck! With thanks

Read More

Answer

All 3 monoclonal antibodies are suitable for IHC. The only difference between the 3 is that that ab8319 is an IgG2a, ab8320 is an IgG2b and ab8321 is an IgM. Whichever one you choose, you will need a suitable secondary raised against the mouse subclass of immunoglobulin that you have chosen. For instance, if you choose ab8321, you will need something like a goat anti mouse IgM. I will send you a copy of my antigen retrieval protocol.

Read More

Answer

I have tried to obtain an IHC protocol specific to this antibody but am afraid that I cannot find any more information other than that which already appears on the datasheet. We suspect that antigen retreaval will be necessary and recommend that you try a pressure cooking method first - I shall forward a suggested general protocol. With regard to incubation time, 1hr at RT should be sufficient, but for better staining with a reduced background, we suggest you also try an incubation at 4 deg C O/N.

Read More

Answer

This antibody has been used extensively on ff paraffin sections. We recommend supernatants or lysates of 2 x 106 endotoxin-stimulated human PBMC cells as a +ve control.

Read More

31-38 of 38 Abreviews or Q&A

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