Recombinant Anti-GSDMD antibody [EPR19828] (ab209845)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR19828] to GSDMD
- Suitable for: WB, IP
- Knockout validated
- Reacts with: Mouse
Related conjugates and formulations
Overview
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Product name
Anti-GSDMD antibody [EPR19828]
See all GSDMD primary antibodies -
Description
Rabbit monoclonal [EPR19828] to GSDMD -
Host species
Rabbit -
Specificity
In our hands this product works well in western blot using mouse cell lines but not tissue samples.
We recommend ab219800 as an alternative for low expression samples.
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Tested applications
Suitable for: WB, IPmore details -
Species reactivity
Reacts with: Mouse -
Immunogen
Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.
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Positive control
- WB: RAW 264.7 cell line with ectopic expression of Apoptosis-associated speck-like protein, untreated and primed with 1 µg/mL lipopolysaccharides for 4 h followed by 10 µM nigericin treatment under serum starved conditions for 2 h, whole cell lysates; J774A.1. Mouse liver lysate IP: RAW 264.7-derived cells with ectopic expression of ASC, untreated and primed with 1 µg/mL lipopolysaccharides (LPS) for 4 h followed by 10 µM nigericin treatment under serum starved conditions for 2 h, whole cell lysates.
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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: 7.2
Preservative: 0.01% Sodium azide
Constituents: 59% PBS, 40% Glycerol, 0.05% BSA -
Concentration information loading...
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Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
EPR19828 -
Isotype
IgG -
Research areas
Associated products
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Alternative Versions
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Compatible Secondaries
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Isotype control
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Recombinant Protein
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Related Products
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab209845 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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WB | (2) |
1/1000. Detects a band of approximately 53, 32 kDa (predicted molecular weight: 53 kDa).
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IP |
1/30.
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Notes |
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WB
1/1000. Detects a band of approximately 53, 32 kDa (predicted molecular weight: 53 kDa). |
IP
1/30. |
Target
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Sequence similarities
Belongs to the gasdermin family. - Information by UniProt
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Database links
- Entrez Gene: 69146 Mouse
- SwissProt: Q9D8T2 Mouse
- Unigene: 27385 Mouse
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Alternative names
- 1810036L03Rik antibody
- DF 5L antibody
- DF5L antibody
see all
Images
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All lanes : Anti-GSDMD antibody [EPR19828] (ab209845) at 1/1000 dilution (ab219800)
Lane 1 : RAW264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysate, Batch 1
Lane 2 : Mouse liver lysate
Lane 3 : NIH/3T3 (mouse embyro fibroblast cell line) whole cell lysate
Lane 4 : RAW264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysate, Batch 2
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Predicted band size: 53 kDa
Observed band size: 53 kDaBlocking buffer and concentration: 5% NFDM/TBST
Diluting buffer and concentration: 5% NFDM/TBST
Exposure time: ab209845 - 120 seconds, ab219800 - 20 seconds
Compared with ab209845, ab219800 has a higher affinity in positive samples.
The MW observed is consistent with the literature: PMID 26375003.
The 43 kDa band is likely to be a GSDMD cleavage product by caspase-3 at GSDMD Asp88 (PMID: 30902848, PMID: 28392147, PMID: 30135078).
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Lanes 1-3 : Anti-GSDMD antibody [EPR19828] (ab209845) at 1/1000 dilution
Lanes 4-6 : Anti-GSDMD antibody [EPR20859] (ab219800) at 1/1000 dilution
Lanes 1 & 4 : Raw264.7 (Mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysates
Lanes 2 & 5 : Mouse liver tissue lysate
Lanes 3 & 6 : Mouse spinal cord tissue lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Predicted band size: 53 kDa
Observed band size: 53 kDa
Exposure time: 3 minutesBlocking/Diluting buffer and concentration 5% NFDM/TBST.
Ab209845 work best with cell lysates.
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All lanes : Anti-GSDMD antibody [EPR19828] (ab209845) at 1/1000 dilution
Lane 1 : RAW 264.7 (Mouse macrophage cell line transformed with Abelson murine leukemia virus)-derived cell line with ectopic expression of ASC (Apoptosis-associated speck-like protein), whole cell lysate
Lane 2 : RAW 264.7 (Mouse macrophage cell line transformed with Abelson murine leukemia virus)-derived cell line with ectopic expression of ASC primed with 1 µg/mL lipopolysaccharides (LPS) for 4 h followed by 10 µM nigericin treatment under serum starved conditions for 2 h, whole cell lysate
Lane 3 : Gasdermin-D knockout RAW 264.7 (Mouse macrophage cell line transformed with Abelson murine leukemia virus)-derived cell line with ectopic expression of ASC 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
Predicted band size: 53 kDa
Observed band size: 32,53 kDa why is the actual band size different from the predicted?
Exposure time: 1 minuteBlocking/Dilution buffer: 5% NFDM/TBST.
As a response to LPS stimulation and nigericin treatment, Gasdermin-D is cleaved and Gasdermin-D N-terminal form is detected at 32kDa. Details of RAW264.7-derived cells with ectopic expression of ASC are described in the literature: PMID 26611636
The MW observed is consistent with the literature: PMID 26375003.
The cells were kindly provided by our collaborator Dr. Jiahuai Han, Xiamen University.
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Lane 1 : Anti-GSDMD antibody [EPR19828] (ab209845) at 1/1000 dilution
Lane 2 : Anti-GSDMD antibody [EPR20859] (ab219800) at 1/1000 dilution
All lanes : J774A.1 (Mouse reticulum cell sarcoma monocyte macrophage) whole cell lysates
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution
Predicted band size: 53 kDa
Observed band size: 53 kDa
Exposure time: 3 minutesDiluting/Blocking buffer and concentration 5% NFDM/TBST
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Gasdermin-D was immunoprecipitated from 0.35 mg of RAW 264.7 (Mouse macrophage cell line transformed with Abelson murine leukemia virus)-derived cells with ectopic expression of ASC were primed with 1μg/mL lipopolysaccharides (LPS) for 4 h followed by 10 μM nigericin treatment under serum starved conditions for 2 h, whole cell lysate with ab209845 at 1/30 dilution.
Western blot was performed from the immunoprecipitate using ab209845 at 1/1000 dilution.
VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 3 minutes.
As a response to LPS stimulation and nigericin treatment, Gasdermin-D is cleaved and Gasdermin-D N-terminal form is detected at 32kDa. Details of RAW 264.7-derived cells with ectopic expression of ASC are described in the literature: PMID 26611636.
The cells were kindly provided by our collaborator Dr. Jiahuai Han, Xiamen University.
Note: The antibody has better affinity to full length Gasdermin-D.
All lanes :
Lane 1 : RAW 264.7-derived cells with ectopic expression of ASC were primed with 1µg/mL lipopolysaccharides (LPS) for 4 h followed by 10 µM nigericin treatment under serum starved conditions for 2 h at 10 µg
Lane 2 : ab209845 IP in RAW 264.7-derived cells with ectopic expression of ASC were primed with 1µg/mL lipopolysaccharides (LPS) for 4 h followed by 10 µM nigericin treatment under serum starved conditions for 2 h whole cell lysate at 10 µg
Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab209845 in RAW 264.7-derived cells with ectopic expression of ASC were primed with 1µg/mL lipopolysaccharides (LPS) for 4 h followed by 10 µM nigericin treatment under serum starved conditions for 2 h whole cell lysate -
Different batches of ab209845 were tested on:
1: RAW 264.7 (Mouse macrophage cell line transformed with Abelson murine leukemia virus) -derived cell line with ectopic expression of ASC primed with 1 µg/mL lipopolysaccharides (LPS) for 4 h followed by 10 uM nigericin treatment under serum starved conditions for 2 h, whole cell lysate at 5.2 µg/ml.
2: RAW 264.7 (Mouse macrophage cell line transformed with Abelson murine leukemia virus) -derived cell line with ectopic expression of ASC (Apoptosis-associated speck-like protein), whole cell lysate at 5.2 µg/ml.
15 µg of lysate was loaded in each lane. Bands observed at 32,53 kDa.
Protocols
Datasheets and documents
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SDS download
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Datasheet download
Certificate of Compliance
References (195)
ab209845 has been referenced in 195 publications.
- Li Y et al. Bacterial outer membrane vesicles-based therapeutic platform eradicates triple-negative breast tumor by combinational photodynamic/chemo-/immunotherapy. Bioact Mater 20:548-560 (2023). PubMed: 35846843
- Zhang LM et al. STING mediates neuroinflammatory response by activating NLRP3-related pyroptosis in severe traumatic brain injury. J Neurochem 162:444-462 (2022). PubMed: 35892155
- Feng S et al. Pathogen-selective killing by guanylate-binding proteins as a molecular mechanism leading to inflammasome signaling. Nat Commun 13:4395 (2022). PubMed: 35906252
- Li X et al. Caspase-1 and Gasdermin D Afford the Optimal Targets with Distinct Switching Strategies in NLRP1b Inflammasome-Induced Cell Death. Research (Wash D C) 2022:9838341 (2022). PubMed: 35958114
- Wang X et al. Schisandrin A from Schisandra chinensis Attenuates Ferroptosis and NLRP3 Inflammasome-Mediated Pyroptosis in Diabetic Nephropathy through Mitochondrial Damage by AdipoR1 Ubiquitination. Oxid Med Cell Longev 2022:5411462 (2022). PubMed: 35996380