Validated using a knockout cell lineRecombinantRabMAb

Recombinant Anti-NLRP3 antibody [EPR23094-1] (ab263899)

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Western blot - Anti-NLRP3 antibody [EPR23094-1] (ab263899)
  • Western blot - Anti-NLRP3 antibody [EPR23094-1] (ab263899)
  • Western blot - Anti-NLRP3 antibody [EPR23094-1] (ab263899)
  • Western blot - Anti-NLRP3 antibody [EPR23094-1] (ab263899)
  • Immunoprecipitation - Anti-NLRP3 antibody [EPR23094-1] (ab263899)
  • Immunoprecipitation - Anti-NLRP3 antibody [EPR23094-1] (ab263899)
  • Flow Cytometry - Anti-NLRP3 antibody [EPR23094-1] (ab263899)
  • ELISA - Anti-NLRP3 antibody [EPR23094-1] (ab263899)
  • Anti-NLRP3 antibody [EPR23094-1] (ab263899)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EPR23094-1] to NLRP3
  • Suitable for: ELISA, WB, Flow Cyt, IP
  • Knockout validated
  • Reacts with: Mouse, Rat, Human, Recombinant fragment

Overview

  • Product name

    Anti-NLRP3 antibody [EPR23094-1]
    See all NLRP3 primary antibodies

  • Description

    Rabbit monoclonal [EPR23094-1] to NLRP3

  • Host species

    Rabbit

  • Specificity

    Stimulation may be required to allow detection of the NLRP3 protein in most samples.

  • Tested applications

    Suitable for: ELISA, WB, Flow Cyt, IPmore details
    Unsuitable for: ICC/IF or IHC-P

  • Species reactivity

    Reacts with: Mouse, Rat, Human, Recombinant fragment

  • Immunogen

    Recombinant fragment. This information is proprietary to Abcam and/or its suppliers.

  • Positive control

    • WB: THP-1 and J774A.1 whole cell lysate. LPS treated RAW 264.7 whole cell lysate. Rat spleen lysate. Mouse Wt BMDCs treated with 50 ng/ml LPS, 3h; Human THP1 Wt cells treated with 200 ng/ml LPS, 3h. RAW 264.7 (Mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysate. IP: RAW 264.7 and THP-1 whole cell lysate. Flow Cyt: THP-1 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
    For more information see here.

    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

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab263899 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
ELISA
Use at an assay dependent concentration.
WB (2)
1/1000. Predicted molecular weight: 118 kDa.

Stimulation may be required to allow detection of the NLRP3 protein in most samples.

There may be multiple bands of NLRP3 in WB assay due to different isoforms and PTMs.
Flow Cyt
1/50.

 

 
IP
1/30.

 

 
Notes
ELISA
Use at an assay dependent concentration.
WB
1/1000. Predicted molecular weight: 118 kDa.

Stimulation may be required to allow detection of the NLRP3 protein in most samples.

There may be multiple bands of NLRP3 in WB assay due to different isoforms and PTMs.
Flow Cyt
1/50.

 

 
IP
1/30.

 

 
  • Application notes
    Is unsuitable for ICC/IF or IHC-P.

    • Target

    • Function

      May function as an inducer of apoptosis. Interacts selectively with ASC and this complex may function as an upstream activator of NF-kappa-B signaling. Inhibits TNF-alpha induced activation and nuclear translocation of RELA/NF-KB p65. Also inhibits transcriptional activity of RELA. Activates caspase-1 in response to a number of triggers including bacterial or viral infection which leads to processing and release of IL1B and IL18.

    • Tissue specificity

      Expressed in blood leukocytes. Strongly expressed in polymorphonuclear cells and osteoblasts. Undetectable or expressed at a lower magnitude in B- and T-lymphoblasts, respectively. High level of expression detected in chondrocytes. Detected in non-keratinizing epithelia of oropharynx, esophagus and ectocervix and in the urothelial layer of the bladder.

    • Involvement in disease

      Defects in NLRP3 are the cause of familial cold autoinflammatory syndrome type 1 (FCAS1) [MIM:120100]; also known as familial cold urticaria. FCAS are rare autosomal dominant systemic inflammatory diseases characterized by episodes of rash, arthralgia, fever and conjunctivitis after generalized exposure to cold.
      Defects in NLRP3 are a cause of Muckle-Wells syndrome (MWS) [MIM:191900]; also known as urticaria-deafness-amyloidosis syndrome. MWS is a hereditary periodic fever syndrome characterized by fever, chronic recurrent urticaria, arthralgias, progressive sensorineural deafness, and reactive renal amyloidosis. The disease may be severe if generalized amyloidosis occurs.
      Defects in NLRP3 are the cause of chronic infantile neurologic cutaneous and articular syndrome (CINCA) [MIM:607115]; also known as neonatal onset multisystem inflammatory disease (NOMID). CINCA is a rare congenital inflammatory disorder characterized by a triad of neonatal onset of cutaneous symptoms, chronic meningitis and joint manifestations with recurrent fever and inflammation.

    • Sequence similarities

      Belongs to the NLRP family.
      Contains 1 DAPIN domain.
      Contains 9 LRR (leucine-rich) repeats.
      Contains 1 NACHT domain.

    • Cellular localization

      Cytoplasm.
    • Information by UniProt

    • Database links

      • Alternative names

        • AGTAVPRL antibody
        • AII/AVP antibody
        • Angiotensin/vasopressin receptor AII/AVP like antibody
        • Angiotensin/vasopressin receptor AII/AVP-like antibody
        • C1orf7 antibody
        • Caterpiller protein 1.1 antibody
        • CIAS 1 antibody
        • CIAS1 antibody
        • CLR1.1 antibody
        • Cold autoinflammatory syndrome 1 antibody
        • Cold autoinflammatory syndrome 1 protein antibody
        • Cryopyrin antibody
        • Familial cold autoinflammatory syndrome antibody
        • FCAS antibody
        • FCU antibody
        • LRR and PYD domains-containing protein 3 antibody
        • Muckle-Wells syndrome antibody
        • MWS antibody
        • NACHT antibody
        • NACHT LRR and PYD containing protein 3 antibody
        • NALP 3 antibody
        • NALP3 antibody
        • NALP3_HUMAN antibody
        • NLR family pyrin domain containing 3 antibody
        • NLRP3 antibody
        • PYPAF 1 antibody
        • PYPAF1 antibody
        • PYRIN containing APAF1 like protein 1 antibody
        • PYRIN-containing APAF1-like protein 1 antibody
        see all

      Images

      • Western blot - Anti-NLRP3 antibody [EPR23094-1] (ab263899)
        Western blot - Anti-NLRP3 antibody [EPR23094-1] (ab263899)This image is courtesy of Benedikt Saller, from Prof. Dr. Olaf Groß’ lab at the University of Freiburg.
        All lanes : Anti-NLRP3 antibody [EPR23094-1] (ab263899) at 1/1000 dilution

        Lane 1 : Mouse Wt BMDCs treated with 50 ng/ml LPS, 3h.
        Lane 2 : Mouse NLRP3-/- BMDCs treated with 50 ng/ml LPS, 3h.
        Lane 3 : Human THP1 Wt cells treated with 200 ng/ml LPS, 3h.

        Lysates/proteins at 25000 cells per lane.

        Secondary
        All lanes : Goat anti-Rabbit HRP at 1/5000 dilution

        Developed using the ECL technique.

        Predicted band size: 118 kDa


        Exposure time: 10 seconds


        Blocking step: 2% SMP in PBS/T for 60 minutes at RT.

      • Western blot - Anti-NLRP3 antibody [EPR23094-1] (ab263899)
        Western blot - Anti-NLRP3 antibody [EPR23094-1] (ab263899)
        All lanes : Anti-NLRP3 antibody [EPR23094-1] (ab263899) at 1/1000 dilution

        Lane 1 : THP-1 (human monocytic leukemia monocyte) whole cell lysate at 20 µg
        Lane 2 : Jurkat (human t cell leukemia t lymphocyte) whole cell lysate at 20 µg
        Lane 3 : Untreated RAW 264.7 (mouse abelson murine leukemia virus-induced tumor macrophage) whole cell lysate at 10 µg
        Lane 4 : RAW 264.7 treated with 10 µg/ml LPS for 8 hours, whole cell lysate at 10 µg
        Lane 5 : J774A.1 (mouse reticulum cell sarcoma monocyte macrophage) whole cell lysate at 20 µg

        Secondary
        All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

        Predicted band size: 118 kDa



        The molecular weight/expression profile observed is consistent with what has been described in the literature (PMID: 26939933, 30315268).

        Note: The band around 75kDa is the short form of NLRP3 (NLRP3s).

        Low expression cell line: Jurkat (PMID: 19767079).

        Exposure times: Lane 1: 48 secs; Lane 2: 3 min; Lanes 3-4: 1 sec; Lane 5: 6 secs.

      • Western blot - Anti-NLRP3 antibody [EPR23094-1] (ab263899)
        Western blot - Anti-NLRP3 antibody [EPR23094-1] (ab263899)
        All lanes : Anti-NLRP3 antibody [EPR23094-1] (ab263899) at 1/1000 dilution

        Lane 1 : C6 (Rat glial tumor glial cell) whole cell lysate
        Lane 2 : RAW 264.7 (Mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysate
        Lane 3 : PC-12 (Rat adrenal gland pheochromocytoma) whole cell lysate
        Lane 4 : NIH/3T3 (Mouse embryonic fibroblast) whole cell lysate

        Lysates/proteins at 10 µg per lane.

        Secondary
        All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/10000 dilution

        Predicted band size: 118 kDa
        Observed band size: 118 kDa


        Exposure time: 15 seconds


        Stimulation is required to allow detection of the NLRP3 protein in C6, PC-12 and NIH/3T3 cell lines.

      • Western blot - Anti-NLRP3 antibody [EPR23094-1] (ab263899)
        Western blot - Anti-NLRP3 antibody [EPR23094-1] (ab263899)
        Anti-NLRP3 antibody [EPR23094-1] (ab263899) at 1/1000 dilution + Rat spleen lysate at 10 µg

        Secondary
        Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

        Predicted band size: 118 kDa


        Exposure time: 3 minutes


        The molecular weight/expression profile observed is consistent with what has been described in the literature (PMID: 26939933, 30315268).

      • Immunoprecipitation - Anti-NLRP3 antibody [EPR23094-1] (ab263899)
        Immunoprecipitation - Anti-NLRP3 antibody [EPR23094-1] (ab263899)

        NLRP3 was immunoprecipitated from 0.35 mg RAW 264.7 (Mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysate 10µg with ab263899 at 1/30 dilution (2µg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab263899 at 1/500 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366) was used at 1/1000 dilution.

        Lane 1: RAW 264.7 whole cell lysate 10µg

        Lane 2: ab263899 IP in RAW 264.7 whole cell lysate

        Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab263899 in RAW 264.7 whole cell lysate

        Blocking and dilution buffer and concentration: 5% NFDM/TBST.

        Exposure time: 10 seconds.

         

      • Immunoprecipitation - Anti-NLRP3 antibody [EPR23094-1] (ab263899)
        Immunoprecipitation - Anti-NLRP3 antibody [EPR23094-1] (ab263899)

        NLRP3 was immunoprecipitated from 0.35 mg THP-1 (Human monocytic leukemia monocyte) whole cell lysate 10µg with ab263899 at 1/30 dilution (2µg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab263899 at 1/500 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366) was used at 1/1000 dilution.

        Lane 1: THP-1 whole cell lysate 10µg

        Lane 2: ab263899 IP in THP-1 whole cell lysate

        Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab263899 in THP-1 whole cell lysate

        Blocking and dilution buffer and concentration: 5% NFDM/TBST.

        Exposure time: 30 seconds.

         

      • Flow Cytometry - Anti-NLRP3 antibody [EPR23094-1] (ab263899)
        Flow Cytometry - Anti-NLRP3 antibody [EPR23094-1] (ab263899)

        Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized THP-1 (Human monocytic leukemia monocyte) cells labeling NLRP3 with ab263899 at 1/50 (Red) compared with a Rabbit monoclonal IgG (ab172730) / Black isotype control and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (Blue). A Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) at 1/2000 dilution was used as the secondary antibody.

      • ELISA - Anti-NLRP3 antibody [EPR23094-1] (ab263899)
        ELISA - Anti-NLRP3 antibody [EPR23094-1] (ab263899)
        ELISA analysis of NLRP3 recombinant protein at 1000 ng/mL with ab263899. An Alkaline Phosphatase-conjugated AffiniPure Goat Anti-Rabbit IgG (H+L) at 1/2500 dilution was used as the secondary antibody.
      • Anti-NLRP3 antibody [EPR23094-1] (ab263899)
        Anti-NLRP3 antibody [EPR23094-1] (ab263899)

      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.

      Click here to view the general protocols

      References (18)

      Publishing research using ab263899? Please let us know so that we can cite the reference in this datasheet.

      ab263899 has been referenced in 18 publications.

      • Cai L  et al. Glucocorticoid receptors involved in melatonin inhibiting cell apoptosis and NLRP3 inflammasome activation caused by bacterial toxin pyocyanin in colon. Free Radic Biol Med 162:478-489 (2021). WB ; Human . PubMed: 33189867
      • Yang Y  et al. Impairment of sirtuin 1-mediated DNA repair is involved in bisphenol A-induced aggravation of macrophage inflammation and atherosclerosis. Chemosphere 265:128997 (2021). WB ; Mouse . PubMed: 33239236
      • Guo H  et al. Carthamin yellow improves cerebral ischemia-reperfusion injury by attenuating inflammation and ferroptosis in rats. Int J Mol Med 47:1 (2021). WB ; Rat . PubMed: 33576458
      • Li Z  et al. Long noncoding RNA MEG3 contributes to dysfunction of brain microvascular endothelial cells after intracerebral hemorrhage by regulating the miR-1930-5p/Mllt1 axis. Brain Res Bull 166:1-11 (2021). WB ; Mouse . PubMed: 33127454
      • Chai GR  et al. Quercetin protects against diabetic retinopathy in rats by inducing heme oxygenase-1 expression. Neural Regen Res 16:1344-1350 (2021). WB ; Rat . PubMed: 33318415
      View all Publications for this product

      Customer reviews and Q&As

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      1-2 of 2 Abreviews or Q&A

      Western blot abreview for Anti-NLRP3 antibody [EPR23094-1]

       Excellent
      Abreviews
      abreview image
      Application
      Western blot
      Sample
      Mouse Cell lysate - whole cell (Lysates of BMDCs and Thp1)
      Gel Running Conditions
      Reduced Denaturing (Continous 12% gel)
      Loading amount
      25000 cells
      Treatment
      LPS (3h, 50 ng/ml for BMDCs or 200 ng/ml for Thp1)
      Specification
      Lysates of BMDCs and Thp1
      Blocking step
      Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 2% · Temperature: 25°C
      Read More

      Mr. Benedikt Saller

      Verified customer

      Submitted Jul 02 2020

      Western blot abreview for Anti-NLRP3 antibody [EPR23094-1]

       Below Average
      Abreviews
      Application
      Western blot
      Sample
      Rat Cell lysate - whole cell (endothelial cells)
      Gel Running Conditions
      Reduced Denaturing (10%)
      Loading amount
      30 µg
      Specification
      endothelial cells
      Blocking step
      Milk as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 23°C
      Read More

      Dr. Dhanush Haspula

      Verified customer

      Submitted May 26 2020

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