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25620
Human Reactive Inflammasome Antibody Sampler Kit II
Primary Antibodies
Antibody Sampler Kit
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Human Reactive Inflammasome Antibody Sampler Kit II #25620

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Human Reactive Inflammasome Antibody Sampler Kit II: Image 1
Simple Western™ analysis of lysates (0.1 mg/mL) from THP-1 cells using NLRP3 (D4D8T) Rabbit mAb #15101. The virtual lane view (left) shows the target band (as indicated) at 1:10 and 1:50 dilutions of primary antibody. The corresponding electropherogram view (right) plots chemiluminescence by molecular weight along the capillary at 1:10 (blue line) and 1:50 (green line) dilutions of primary antibody. This experiment was performed under reducing conditions on the Jess™ Simple Western instrument from ProteinSimple, a BioTechne brand, using the 12-230 kDa separation module.
Human Reactive Inflammasome Antibody Sampler Kit II: Image 2
Western blot analysis of extracts from THP-1 and MUTZ-3 cells using NLRC4 (D5Y8E) Rabbit mAb.
Human Reactive Inflammasome Antibody Sampler Kit II: Image 3
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).
Human Reactive Inflammasome Antibody Sampler Kit II: Image 4
Western blot analysis of extracts from Daudi, KARPAS-299, and L-540 cell lines using AIM2 (D5X7K) Rabbit mAb.
Human Reactive Inflammasome Antibody Sampler Kit II: Image 5
Western blot analysis of extracts from various cell lines using ASC/TMS1 (E1E3I) Rabbit mAb (upper) or β-Actin (D6A8) Rabbit mAb #8457 (lower). As expected, Jurkat cells do not express ASC/TMS1.
Human Reactive Inflammasome Antibody Sampler Kit II: Image 6
Western blot analysis of extracts from mouse bone marrow-derived dendritic cells (BMDC) and various cell lines using NLRP3 (D4D8T) Rabbit mAb (upper) and β-Actin (D6A8) Rabbit mAb #8457 (lower).
Human Reactive Inflammasome Antibody Sampler Kit II: Image 7
Western blot analysis of extracts from THP-1 and NK-92 cells using Caspase-1 (D7F10) Rabbit mAb.
Human Reactive Inflammasome Antibody Sampler Kit II: Image 8
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).
Human Reactive Inflammasome Antibody Sampler Kit II: Image 9
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.
Human Reactive Inflammasome Antibody Sampler Kit II: Image 10
Western blot analysis of recombinant Human Interleukin-1β (hIL-1β) #8900 using Cleaved-IL-1β (Asp116) (D3A3Z) Rabbit mAb.
Human Reactive Inflammasome Antibody Sampler Kit II: Image 11
Western blot analysis of extracts from 293T cells, mock transfected (-) or transfected with a construct expressing full-length human NLRC4 (hNLRC4; +), using NLRC4 (D5Y8E) Rabbit mAb.
Human Reactive Inflammasome Antibody Sampler Kit II: Image 12
Western blot analysis of recombinant Human Interleukin-1β (hIL-1β) #8900 using IL-1β (D3U3E) Rabbit mAb.
Human Reactive Inflammasome Antibody Sampler Kit II: Image 13
Western blot analysis of extracts from HL-60 cells, serum-starved and either untreated (-) or treated overnight with Human Interferon-γ (hIFN-γ) #8901 (100 ng/ml; +), using AIM2 (D5X7K) Rabbit mAb (upper) or β-Actin (D6A8) Rabbit mAb #8457 (lower).
Human Reactive Inflammasome Antibody Sampler Kit II: Image 14
Immunoprecipitation of ASC/TMS1 from THP-1 cell extracts using Rabbit (D1AE) mAb XP® Isotype Control #3900 (lane 2) or ASC/TMS1 (E1E3I) Rabbit mAb (lane 3). Lane 1 is 10% input. Western blot was performed using ASC/TMS1 (E1E3I) Rabbit mAb.
Human Reactive Inflammasome Antibody Sampler Kit II: Image 15
Immunoprecipitation of NLRP3 from J774A.1 cell extracts using Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (lane 2) or NLRP3 (D4D8T) Rabbit mAb (lane 3). Lane 1 is 10% input. Western blot analysis was performed using NLRP3 (D4D8T) Rabbit mAb.
Human Reactive Inflammasome Antibody Sampler Kit II: Image 16
Western blot analysis of extracts from COS cells, untransfected or transfected with human caspase-1, using Caspase-1 (D7F10) Rabbit mAb.
Human Reactive Inflammasome Antibody Sampler Kit II: Image 17
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 ug/ml, 8 hours), using Cleaved Caspase-1 (Asp297) (D57A2) Rabbit mAb.
Human Reactive Inflammasome Antibody Sampler Kit II: Image 18
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.
Human Reactive Inflammasome Antibody Sampler Kit II: Image 19
Immunoprecipitation of NLRC4 from MUTZ-3 cell extracts using Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (lane 2) or NLRC4 (D5Y8E) Rabbit mAb (lane 3). Lane 1 is 10% input. Western blot analysis was performed using NLRC4 (D5Y8E) Rabbit mAb.
Human Reactive Inflammasome Antibody Sampler Kit II: Image 20
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).
Human Reactive Inflammasome Antibody Sampler Kit II: Image 21
Western blot analysis of extracts from 293T cells, mock transfected (-) or transfected with a construct expressing Myc-DDK tagged human AIM2 (hAIM2-Myc/DDK; +), using AIM2 (D5X7K) Rabbit mAb (upper) and Myc-Tag (71D10) Rabbit mAb #2278 (lower).
Human Reactive Inflammasome Antibody Sampler Kit II: Image 22
Immunohistochemical analysis of paraffin-embedded human ductal breast carcinoma using ASC/TMS1 (E1E3I) Rabbit mAb.
Human Reactive Inflammasome Antibody Sampler Kit II: Image 23
Flow cytometric analysis of THP-1, untreated (blue, negative) or treated with LPS #14011 (100 ng/ml, 3 hr; green, positive) using IL-1β (D3U3E) Rabbit mAb (solid lines) or concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (dashed lines). Anti-rabbit IgG (H+L), F(ab')₂ Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.
Human Reactive Inflammasome Antibody Sampler Kit II: Image 24
Immunoprecipitation of AIM2 from HL-60 cell extracts treated with Human Interferon-γ (hIFN-γ) #8901 (100 ng/ml, overnight) using Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (lane 2) or AIM2 (D5X7K) Rabbit mAb (lane 3). Lane 1 is 10% input. Western blot was performed using AIM2 (D5X7K) Rabbit mAb.
Human Reactive Inflammasome Antibody Sampler Kit II: Image 25
Immunohistochemical analysis of paraffin-embedded THP-1 cell pellet (left, positive) or Jurkat cell pellet (right, negative) using ASC/TMS1 (E1E3I) Rabbit mAb.
Human Reactive Inflammasome Antibody Sampler Kit II: Image 26
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).
Human Reactive Inflammasome Antibody Sampler Kit II: Image 27
Immunohistochemical analysis of paraffin-embedded human colon carcinoma using ASC/TMS1 (E1E3I) Rabbit mAb (left) compared to concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (right).
Human Reactive Inflammasome Antibody Sampler Kit II: Image 28
Immunohistochemical analysis of paraffin-embedded human renal cell carcinoma using ASC/TMS1 (E1E3I) Rabbit mAb.
Human Reactive Inflammasome Antibody Sampler Kit II: Image 29
Immunohistochemical analysis of paraffin-embedded human T-cell lymphoma using ASC/TMS1 (E1E3I) Rabbit mAb.
Human Reactive Inflammasome Antibody Sampler Kit II: Image 30
Immunohistochemical analysis of paraffin-embedded human spleen using ASC/TMS1 (E1E3I) Rabbit mAb.
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To Purchase # 25620
Cat. # Size Qty. Price
25620T		 1 Kit  (8 x 20 microliters)
$ 645

Custom Formulation

Product Includes Quantity Applications Reactivity MW(kDa) Isotype
NLRP3 (D4D8T) Rabbit mAb 15101 20 µl
  • WB
  • IP
 H M 110 Rabbit IgG
AIM2 (D5X7K) Rabbit mAb 12948 20 µl
  • WB
  • IP
 H 40 Rabbit IgG
NLRC4 (D5Y8E) Rabbit mAb 12421 20 µl
  • WB
  • IP
 H 110 Rabbit IgG
Caspase-1 (D7F10) Rabbit mAb 3866 20 µl
  • WB
  • IP
 H 48, 20 Rabbit IgG
ASC/TMS1 (E1E3I) Rabbit mAb 13833 20 µl
  • WB
  • IP
  • IHC
 H 22, 19, 15 Rabbit IgG
Cleaved Caspase-1 (Asp297) (D57A2) Rabbit mAb 4199 20 µl
  • WB
  • IP
 H 20, 22 Rabbit IgG
Cleaved-IL-1β (Asp116) (D3A3Z) Rabbit mAb 83186 20 µl
  • WB
  • IF
 H 17 Rabbit IgG
IL-1β (D3U3E) Rabbit mAb 12703 20 µl
  • WB
  • IF
  • F
 H 17, 31 Rabbit IgG
Anti-rabbit IgG, HRP-linked Antibody 7074 100 µl
  • WB
 Rab Goat 

Product Description

The Human Reactive Inflammasome Antibody Sampler Kit II provides an economical means of detecting multiple inflammasome components. The kit contains enough primary antibodies to perform at least two western blot experiments.

Specificity / Sensitivity

Each antibody in the Human Reactive Inflammasome Antibody Sampler Kit II detects endogenous levels of its target protein. AIM2 (D5X7K) Rabbit mAb detects a 22 kDa band of unknown origin in some cell lines. Caspase-1 (D7F10) Rabbit mAb detects endogenous levels of full-length human Caspase-1; the activated p20 subunit was detected by overexpression. ASC/TMS1 (E1E3I) Rabbit mAb can detect three known isoforms of ASC/TMS1. Cleaved Caspase-1 (Asp297) (D57A2) Rabbit mAb detects endogenous levels of the p20 subunit of human caspase-1 only upon cleavage at Asp297. Cleaved-IL-1β (Asp116) (D3A3Z) Rabbit mAb recognizes endogenous levels of mature IL-1β protein only when cleaved at Asp116. IL-1β (D3U3E) Rabbit mAb is not able to detect endogenous levels of mature IL-1β. It can detect up to 100 pg of recombinant mature IL-1β.

Source / Purification

Monoclonal antibodies are produced by immunizing animals with recombinant human IL-1β protein or with synthetic peptides corresponding to residues adjacent to Asp297 human caspase-1, residues adjacent to Asp116 of human IL-1β, residues surrounding Ala306 of mouse NLRP3, Lys93 of human AIM2, Leu942 of human NLRC4, and the carboxy terminus of human ASC/TMS1 isoform 1.

Background

The innate immune system works as the first line of defense in protection from pathogenic microbes and host-derived signals of cellular distress. One way in which these “danger” signals trigger inflammation is through activation of inflammasomes, which are multiprotein complexes that assemble in the cytosol after exposure to pathogen-associated molecular patterns (PAMPs) or danger-associated molecular patterns (DAMPs) and result in the activation of caspase-1 and subsequent cleavage of proinflammatory cytokines IL-1β and IL-18 (Reviewed in 1-6). Inflammasome complexes typically consist of a cytosolic pattern recognition receptor (PRR; a nucleotide-binding domain and leucine-rich-repeat [NLR] or AIM2-like receptor [ALR] family member), an adaptor protein (ASC/TMS1), and pro-caspase-1. A number of distinct inflammasome complexes have been identified, each with a unique PRR and activation triggers. The best characterized is the NLRP3 complex, which contains NLRP3, ASC/TMS1, and pro-caspase-1. The NLRP3 inflammasome is activated in a two-step process. First, NF-κB signaling is induced through PAMP- or DAMP-mediated activation of TLR4 or TNFR, resulting in increased expression of NLRP3, pro-IL-1β, and pro-IL-18 (priming step, signal 1). Next, indirect activation of NLRP3 occurs by a multitude of signals (whole pathogens, PAMPs/DAMPs, potassium efflux, lysosomal-damaging environmental factors [uric acid, silica, alum] and endogenous factors [amyloid-β, cholesterol crystals], and mitochondrial damage), leading to complex assembly and activation of caspase-1 (signal 2). The complex inflammasome structure is built via domain interactions among the protein components. Other inflammasomes are activated by more direct means: double-stranded DNA activates the AIM2 complex, anthrax toxin activates NLRP1, and bacterial flagellin activates NLRC4. Activated caspase-1 induces secretion of proinflammatory cytokines IL-1β and -18, but also regulates metabolic enzyme expression, phagosome maturation, vasodilation, and pyroptosis, an inflammatory programmed cell death. Inflammasome signaling contributes to the onset of a number of diseases, including atherosclerosis, type II diabetes, Alzheimer’s disease, and autoimmune disorders.
  1. Broz, P. and Dixit, V.M. (2016) Nat Rev Immunol 16, 407-20.
  2. Guo, H. et al. (2015) Nat Med 21, 677-87.
  3. Jo, E.K. et al. (2016) Cell Mol Immunol 13, 148-59.
  4. Rathinam, V.A. and Fitzgerald, K.A. (2016) Cell 165, 792-800.
  5. Shao, B.Z. et al. (2015) Front Pharmacol 6, 262.
  6. Schroder, K. and Tschopp, J. (2010) Cell 140, 821-32.

Pathways

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For Research Use Only. Not For Use In Diagnostic Procedures.
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