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43811
Pyroptosis Antibody Sampler Kit
Primary Antibodies
Antibody Sampler Kit

Pyroptosis Antibody Sampler Kit #43811

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Pyroptosis Antibody Sampler Kit: Image 1

Immunoprecipitation of Gasdermin D protein from THP-1 cell extracts. Lane 1 is 10% input, lane 2 is Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is Gasdermin D (E8G3F) Rabbit mAb. Western blot analysis was performed using Gasdermin D (E8G3F) Rabbit mAb. Mouse Anti-rabbit IgG (Conformation Specific) (L27A9) mAb (HRP Conjugate) #5127 was used as a secondary antibody.

Pyroptosis Antibody Sampler Kit: Image 2

Immunohistochemical analysis of paraffin-embedded human ductal breast carcinoma using Cleaved Gasdermin D (Asp275) (E7H9G) Rabbit mAb.

Pyroptosis Antibody Sampler Kit: Image 3

Western blot analysis of extracts from THP-1 and NK-92 cells using Caspase-1 (D7F10) Rabbit mAb.

Pyroptosis Antibody Sampler Kit: Image 4

Western blot analysis of extracts from COS-7 cells, untransfected () or transfected with construct overexpressing human caspase-1 (+), using Cleaved Caspase-1 (Asp297) (D57A2) Rabbit mAb (upper) or Caspase-1 (D7F10) Rabbit mAb #3866 (lower).

Pyroptosis Antibody Sampler Kit: Image 5

Flow cytometric analysis of THP-1 cells, untreated (blue) or LPS-treated (100 ng/ml, 3 hr; green), using IL-1β (D3U3E) Rabbit mAb. Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 647 Conjugate) #4414 was used as a secondary antibody.

Pyroptosis Antibody Sampler Kit: Image 6

Confocal immunofluorescent analysis of THP-1 cells, differentiated with TPA #4174 (80 nM, 24 hr) and subsequently treated with (right) or without (left) Lipopolysaccharides (LPS) #14011 (1 μg/ml, 6 hr), using Cleaved-IL-1β (Asp116) (D3A3Z) Rabbit mAb (green). Actin filaments were labeled with DyLight™ 554 Phalloidin #13054 (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).

Pyroptosis Antibody Sampler Kit: Image 7

Western blot analysis of extracts from KARPAS-299, THP-1 and IM-9 cells using Caspase-4 Antibody.

Pyroptosis Antibody Sampler Kit: Image 8

Western blot analysis of extracts from THP-1 cells differentiated with TPA #4174 (80 nM, overnight) followed by treatment with LPS #14011 (1 μg/ml, indicated times), using Caspase-5 (D3G4W) Rabbit mAb (upper) and β-Actin (D6A8) Rabbit mAb #8457 (lower)

Pyroptosis Antibody Sampler Kit: Image 9

Immunohistochemical analysis of paraffin-embedded mouse lung using HMGB1 (D3E5) Rabbit mAb.

Pyroptosis Antibody Sampler Kit: Image 10

After the primary antibody is bound to the target protein, a complex with HRP-linked secondary antibody is formed. The LumiGLO® is added and emits light during enzyme catalyzed decomposition.

Pyroptosis Antibody Sampler Kit: Image 11

Western blot analysis of extracts from PC-3 cells and PC-3 GSDMD knockout (-/-) cells using Gasdermin D (E8G3F) Rabbit mAb (upper) or GAPDH (D16H11) XP® Rabbit mAb #5174 (lower).

Pyroptosis Antibody Sampler Kit: Image 12

Immunohistochemical analysis of paraffin-embedded human colon carcinoma using Cleaved Gasdermin D (Asp275) (E7H9G) Rabbit mAb in the presence of non-cleaved Gasdermin D peptide (left) or Asp275 cleavage-specific Gasdermin D peptide (right).

Pyroptosis Antibody Sampler Kit: Image 13

Western blot analysis of extracts from COS cells, untransfected or transfected with human caspase-1, using Caspase-1 (D7F10) Rabbit mAb.

Pyroptosis Antibody Sampler Kit: Image 14

Western blot analysis of extracts from the media of THP-1 cells, differentiated with TPA #9905 (80 nM, overnight) followed by treatment with LPS (1 mu-g/ml, 8 hours), using Cleaved Caspase-1 (Asp297) (D57A2) Rabbit mAb.

Pyroptosis Antibody Sampler Kit: Image 15

Confocal immunofluorescent analysis of THP-1 cells, untreated (left) or LPS-treated (500 ng/ml, 2 hr; right), using IL-1β (D3U3E) Rabbit mAb (green). Actin filaments were labeled with DY-554 phalloidin (red).

Pyroptosis Antibody Sampler Kit: Image 16

Immunoprecipitation of Cleaved-IL-1β (Asp116) from extracts of THP-1 cells differentiated with TPA #4147 (80 nM, overnight) followed by treatment with Lipopolysaccharides (LPS) #14011 (1 μg/ml, 6 hr). Lane 1 is 10% input, lane 2 is precipitated with Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is precipitated with Cleaved-IL-1β (Asp116) (D3A3Z) Rabbit mAb. Western blot was performed using Cleaved-IL-1β (Asp116) (D3A3Z) Rabbit mAb.

Pyroptosis Antibody Sampler Kit: Image 17

Western blot analyis of extracts from recombinant, active caspase-1, -4, and -5 using Caspase-4 Antibody.

Pyroptosis Antibody Sampler Kit: Image 18

Western blot analysis of extracts form 293T cells, mock transfected (-) or transfected with constructs expressing full-length human caspase-4 protein (hCasp4; +) or caspase-5 protein (hCasp5; +), using Caspase-5 (D3G4W) Rabbit mAb (upper) and GAPDH (D16H11) XP® Rabbit mAb #5174 (lower).

Pyroptosis Antibody Sampler Kit: Image 19

Immunohistochemical analysis of paraffin-embedded human colon carcioma using HMGB1 (D3E5) Rabbit mAb.

Pyroptosis Antibody Sampler Kit: Image 20

Western blot analysis of extracts from various cell lines using Gasdermin D (E8G3F) Rabbit mAb (upper) or GAPDH (D16H11) XP® Rabbit mAb #5174 (lower).

Pyroptosis Antibody Sampler Kit: Image 21

Immunohistochemical analysis of paraffin-embedded human squamous cell lung carcinoma using Cleaved Gasdermin D (Asp275) (E7H9G) Rabbit mAb.

Pyroptosis Antibody Sampler Kit: Image 22

Western blot analysis of extracts from THP-1 cells, untreated (-) or LPS-treated (100 ng/ml, 3 hr; +), using IL-1β (D3U3E) Rabbit mAb (upper) and β-Actin (D6A8) Rabbit mAb #8457 (lower).

Pyroptosis Antibody Sampler Kit: Image 23

Western blot analysis of recombinant Human Interleukin-1β (hIL-1β) #8900 using Cleaved-IL-1β (Asp116) (D3A3Z) Rabbit mAb.

Pyroptosis Antibody Sampler Kit: Image 24

Immunohistochemical analysis of paraffin-embedded human lung carcioma using HMGB1 (D3E5) Rabbit mAb.

Pyroptosis Antibody Sampler Kit: Image 25

Western blot analysis of extracts from THP-1 cells, differentiated with TPA #4174 (50 ng/ml, overnight) and then treated with LPS #14011 (5 μg/ml, indicated times), using Gasdermin D (E8G3F) Rabbit mAb (upper) or GAPDH (D16H11) XP® Rabbit mAb #5174 (lower).

Pyroptosis Antibody Sampler Kit: Image 26

Immunohistochemical analysis of paraffin-embedded human non-Hodgkin's Lymphoma using Cleaved Gasdermin D (Asp275) (E7H9G) Rabbit mAb.

Pyroptosis Antibody Sampler Kit: Image 27

Western blot analysis of recombinant Human Interleukin-1β (hIL-1β) #8900 using IL-1β (D3U3E) Rabbit mAb.

Pyroptosis Antibody Sampler Kit: Image 28

Western blot analysis of extracts from cells or media collected from THP-1 cells, differentiated with TPA #4147 (80 nM, overnight) and subsequently treated with (+) or without (-) Lipopolysaccharides (LPS) #14011 (1 μg/ml, 6 hr), using Cleaved-IL-1β (Asp116) (D3A3Z) Rabbit mAb.

Pyroptosis Antibody Sampler Kit: Image 29

Immunohistochemical analysis of paraffin-embedded human prostate carcioma using HMGB1 (D3E5) Rabbit mAb.

Pyroptosis Antibody Sampler Kit: Image 30

Immunohistochemical analysis of paraffin-embedded human prostate carcinoma using Cleaved Gasdermin D (Asp275) (E7H9G) Rabbit mAb.

Pyroptosis Antibody Sampler Kit: Image 31

Western blot analysis of extracts from various cell lines using HMGB1 (D3E5) Rabbit mAb.

Pyroptosis Antibody Sampler Kit: Image 32

Immunohistochemical analysis of paraffin-embedded human spleen (left, positive) and skeletal muscle (right, negative) using Cleaved Gasdermin D (Asp275) (E7H9G) Rabbit mAb.

Pyroptosis Antibody Sampler Kit: Image 33

Immunoprecipitation of Cleaved Gasdermin D (Asp725) from THP-1 cells differentiated with TPA #4174 (50 ng/ml, overnight) and then treated with LPS #14011 (5 μg/ml, 6 hr). Lane 1 is 10% input, lane 2 is Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is Cleaved Gasdermin D (Asp275) (E7H9G) Rabbit mAb. Western blot was performed using Cleaved Gasdermin D (Asp275) (E7H9G) Rabbit mAb. Anti-rabbit IgG, HRP-linked Antibody #7074 was used as a secondary antibody.

Pyroptosis Antibody Sampler Kit: Image 34

Western blot analysis of extracts from THP-1 cells, differentiated with TPA #4174 (50 ng/ml, overnight) and then treated with LPS #14011 (5 μg/ml, indicated times), using Cleaved Gasdermin D (Asp275) (E7H9G) Rabbit mAb (upper), total Gasdermin D (L60) Antibody #93709 (middle), or β-Actin (D6A8) Rabbit mAb #8457 (lower).

To Purchase # 43811T
Product # Size Price
43811T
1 Kit  (9 x 20 µl) $ 609

Product Includes Quantity Applications Reactivity MW(kDa) Isotype
Gasdermin D (E8G3F) Rabbit mAb 97558 20 µl
  • WB
  • IP
H 53, 43, 30, 21 Rabbit IgG
Cleaved Gasdermin D (Asp275) (E7H9G) Rabbit mAb 36425 20 µl
  • WB
  • IP
  • IHC
H 30 Rabbit IgG
Caspase-1 (D7F10) Rabbit mAb 3866 20 µl
  • WB
  • IP
H 48, 20 Rabbit IgG
Cleaved Caspase-1 (Asp297) (D57A2) Rabbit mAb 4199 20 µl
  • WB
  • IP
H 20, 22 Rabbit IgG
IL-1β (D3U3E) Rabbit mAb 12703 20 µl
  • WB
  • IF
  • F
H 17, 31 Rabbit IgG
Cleaved-IL-1β (Asp116) (D3A3Z) Rabbit mAb 83186 20 µl
  • WB
  • IP
  • IF
H 17 Rabbit IgG
Caspase-4 Antibody 4450 20 µl
  • WB
H 45 Rabbit 
Caspase-5 (D3G4W) Rabbit mAb 46680 20 µl
  • WB
  • IP
H 50, 44, 35 Rabbit IgG
HMGB1 (D3E5) Rabbit mAb 6893 20 µl
  • WB
  • IHC
H M R Mk 29 Rabbit IgG
Anti-rabbit IgG, HRP-linked Antibody 7074 100 µl
  • WB
Goat 

Product Description

The Pyroptosis Antibody Sampler Kit provides an economical means of detecting proteins that are used as readouts for pyroptosis. The kit includes enough antibodies to perform two western blot experiments with each primary antibody.

Specificity / Sensitivity

Gasdermin D (E8G3F) Rabbit mAb recognizes endogenous levels of total Gasdermin D protein. This antibody recognizes the 30 kDa amino terminal fragment produced during pyroptosis by caspase-1, a 43 kDa fragment produced by caspase-3, as well as a 21 kDa fragment produced by cleavage at both sites. Cleaved Gasdermin D (Asp275) (E7H9G) Rabbit mAb detects the N-terminal fragment of human Gasdermin D protein only when cleaved at Asp275. Caspase-1 (D7F10) Rabbit mAb detects endogenous levels of full length human caspase-1 and the activated p20 subunit by overexpression. Cleaved Caspase-1 (Asp297) (D57A2) Rabbit mAb detects the p20 subunit of human caspase-1 upon cleavage at Asp275. IL-1β (D3U3E) Rabbit mAb can recognize endogenous levels of full-length protein, but does not detect endogenous levels of the mature IL-1β. It can detect up to 100 pg of recombinant mature IL-1β. Cleaved IL-1β (Asp116) (D3A3Z) Rabbit mAb detects endogenous levels of mature IL-1β protein when cleaved at Asp116. Caspase-4 Antibody detects endogenous levels of total caspase-4 and intermediate forms at 40 and 32 kDa. Caspase-5 (D3G4W) Rabbit mAb detects endogenous levels of caspase-5. HMGB1 (D3E5) Rabbit mAb detects endogenous levels of total HMGB1. It does not cross-react with other HMGB proteins, including HMGB2 and HMGB3.

Source / Purification

Monoclonal antibodies are produced by immunizing animals with synthetic peptides corresponding to residues surrounding Trp130 of human Gasdermin D, residues within the p20 subunit of caspase-1, Asp297 of human caspase-1, Asp116 of human IL-1β, Pro154 of human caspase-5, and Ala137 of human HMGB1. IL-1β (D3U3E) Rabbit mAb is produced by immunizing animals with a recombinant IL-1β protein. Polyclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Ile125 of human caspase-4. Polyclonal antibodies are purified by Protein A and peptide affinity chromatography.

Background

Pyroptosis is a regulated pathway of cell death with morphological features of necrosis, including cell swelling, plasma membrane pore formation, and engagement of an inflammatory response with the release of a number of damage-associated molecular patterns (DAMPs), such as HMGB1 and inflammatory cytokines like IL-1β and IL-18 (1,2). Pyroptosis is generally induced in cells of the innate immune system, such as monocytes, macrophages, and dendritic cells in the presence of pathogen-associated molecular patterns (PAMPs) expressed on microbial pathogens or by cell-derived DAMPs. It is induced through assembly of inflammasomes triggering proteolytic activation of caspase-1 which then cleaves inflammatory cytokines like IL-1β and IL-18 to their mature forms (3). A critical feature of pyroptosis is the cleavage of Gasdermin D by caspase-1 and mouse caspase-11 (or human caspase-4/5) (4-6). Upon cleavage, the N-terminal fragment of Gasdermin D oligomerizes to form a pore, allowing secretion of inflammatory DAMPs and cytokines. Canonical inflammasome assembly typically consists of a cytosolic-pattern recognition receptor (PPR; a nucleotide binding domain and leucine-rich repeat [NLR] or AIM2-like family members), an adaptor protein (ASC/TMS1), and pro-caspase-1. Distinct inflammasome complexes can recognize distinct PAMPs and DAMPs to trigger pyroptosis. The best characterized pathway triggered by the NLR, NLRP3, occurs through a two-step process. The first step is a priming signal, NF-κB is activated to induce the expression of a number of inflammasome components including NLRP3, pro-IL-1β, and pro-IL-18. In the second activation step, caspase-1 is activated and Gasdermin D and cytokines are proteolytically activated. In a non-canonical pathway, caspase-4 and caspase-5 can directly trigger Gasdermin D cleavage in monocytes following LPS stimulation (5,7).

  1. Frank, D. and Vince, J.E. (2019) Cell Death Differ 26, 99-114.
  2. Shi, J. et al. (2017) Trends Biochem Sci 42, 245-54.
  3. Malik, A. and Kanneganti, T.D. (2017) J Cell Sci 130, 3955-63.
  4. He, W.T. et al. (2015) Cell Res 25, 1285-98.
  5. Shi, J. et al. (2015) Nature 526, 660-5.
  6. Kayagaki, N. et al. (2015) Nature 526, 666-71.
  7. Viganò, E. et al. (2015) Nat Commun 6, 8761.

Pathways & Proteins

Explore pathways + proteins related to this product.

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