Product Pathways - NF-kB Signaling
Rig-I Pathway Antibody Sampler Kit #8348
|8348S||1 Kit (9 x 40 µl)||---||In Stock||---|
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|Kit Includes||Quantity||Applications||Reactivity||Homology†||MW (kDa)||Isotype|
|MDA-5 (D74E4) Rabbit mAb #5321||40 µl||W, IP||H, M||135||Rabbit IgG|
|Rig-I (D14G6) Rabbit mAb #3743||40 µl||W, IP||H, M, R, Mk||102||Rabbit IgG|
|MAVS Antibody #3993||40 µl||W, IF-IC||H||75, 52||Rabbit|
|IRF-3 (D83B9) Rabbit mAb #4302||40 µl||W, IP||H, M, R, Mk||45-55||Rabbit IgG|
|TBK1/NAK (D1B4) Rabbit mAb #3504||40 µl||W, IP||H, M, R, Mk||84||Rabbit IgG|
|Phospho-TBK1/NAK (Ser172) (D52C2) XP® Rabbit mAb #5483||40 µl||W, IP, IF-IC, F||H||M, R, Mk, X, B, Dg||84||Rabbit IgG|
|Phospho-IRF-3 (Ser396) (4D4G) Rabbit mAb #4947||40 µl||W||H, M||Mk, Pg||45-55||Rabbit IgG|
|IKKε (D61F9) XP® Rabbit mAb #3416||40 µl||W, IP, IF-IC, F||M, R||80||Rabbit|
|IKKε (D20G4) Rabbit mAb #2905||40 µl||W, IP||H||Mk||80||Rabbit IgG|
|Anti-rabbit IgG, HRP-linked Antibody #7074||100 µl||Goat|
†Species predicted to react based on 100% sequence homology.
Applications Key: W=Western Blotting, IP=Immunoprecipitation, IF-IC=Immunofluorescence (Immunocytochemistry), F=Flow Cytometry
Reactivity Key: H=Human, M=Mouse, R=Rat, Mk=Monkey
Western blot analysis of recombinant GST-IKKε and extracts from Ramos and RL7 cells using IKKε (D20G4) Rabbit mAb #2905.
Western blot analysis of extracts from Raw 264.7 and KNRK cells using IKKε (D61F9) XP® Rabbit mAb #3416.
Western blot analysis of extracts from differentiated THP-1 and RAW 264.7 cells, untreated (-) or LPS-treated (1 μg/ml, overnight) (+), using Rig-I (D14G6) Rabbit mAb #3743.
Western blot analysis of extracts from various cell lines using MAVS Antibody #3993.
Western blot analysis of extracts from various cell lines using IRF-3 (D83B9) Rabbit mAb #4302.
Western blot analysis of extracts from differentiated THP-1 cells, untreated (-) or treated with LPS (1 μg/ml, 1 hr) (+), using Phospho-IRF-3 (Ser396) (4D4G) Rabbit mAb #4947 (upper) or IRF-3 (D83B9) Rabbit mAb #4302 (lower).
Western blot anlaysis of extracts from differentiated THP-1 cells, untreated or treated with LPS (1 μg/ml, indicated times), using MDA-5 (D74E4) Rabbit mAb #5321.
The Rig-I Pathway Antibody Sampler Kit provides an economical means to evaluate the activation state and total protein levels of multiple members of the Rig-I pathway including Rig-I, MDA-5, MAVS, IRF-3, TBK1/NAK, and IKKε. The kit includes enough primary antibody to perform four western blot experiments per antibody.
Specificity / Sensitivity
MDA-5 (D74E4) Rabbit mAb, Rig-I (D14G6) Rabbit mAb, MAVS Antibody, IRF-3 (D83B9) Rabbit mAb, TBK1/NAK (D1B4) Rabbit mAb, IKKε (D61F9) XP® Rabbit mAb, and IKKε (D20G4) Rabbit mAb detect endogenous levels of respective total proteins and do not cross-react with other proteins. Bands detected at 52 and 75 kDa by MAVS Antibody correlate with those described by Seth et al. (2005). Phospho-TBK1/NAK (Ser172) (D52C2) XP® Rabbit mAb detects endogenous levels of TBK1/NAK only when phosphorylated at Ser172. Phospho-IRF-3 (Ser396) (4D4G) Rabbit mAb detects endogenous levels of IRF-3 only when phosphorylated at Ser396.
Source / Purification
Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues at the carboxy terminus of human MAVS protein. Polyclonal antibodies are purified by protein A and peptide affinity chromatography. Monoclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Arg470 of human MDA-5 protein, Lys652 of human Rig-I protein, the carboxy terminus of human IRF-3 protein, Ser645 of human TBK1/NAK protein, the carboxy terminus of mouse IKKε protein, or Val345 of human IKKε protein. Activation state monoclonal antibodies are producted by immunizing animals with a synthetic phosphopeptide corresponding to residues surrounding Ser172 of human TBK1/NAK protein or Ser396 of human IRF-3 protein.
Antiviral innate immunity depends on the combination of parallel pathways triggered by virus detecting proteins in the Toll-like receptor (TLR) family and RNA helicases, such as Rig-I (retinoic acid-inducible gene I) and MDA-5 (melanoma differentiation-associated antigen 5), which promote the transcription of type I interferons (IFN) and antiviral enzymes (1-3). TLRs and helicase proteins contain sites that recognize the molecular patterns of different virus types, including DNA, single-stranded RNA (ssRNA), double-stranded RNA (dsRNA), and glycoproteins. These antiviral proteins are found in different cell compartments; TLRs (i.e. TLR3, TLR7, TLR8, and TLR9) are expressed on endosomal membranes and helicases are localized to the cytoplasm. Rig-I expression is induced by retinoic acid, LPS, IFN, and viral infection (4,5). Both Rig-I and MDA-5 share a DExD/H-box helicase domain that detects viral dsRNA and two amino-terminal caspase recruitment domains (CARD) that are required for triggering downstream signaling (4-7). Rig-I binds both dsRNA and viral ssRNA that contains a 5'-triphosphate end not seen in host RNA (8,9). Though structurally related, Rig-I and MDA-5 detect a distinct set of viruses (10,11). The CARD domain of the helicases, which is sufficient to generate signaling and IFN production, is recruited to the CARD domain of the MAVS/VISA/Cardif/IPS-1 mitochondrial protein, which triggers activation of NF-κB, TBK1/IKKε, and IRF-3/IRF-7 (12-15).
- Yoneyama, M. and Fujita, T. (2007) J Biol Chem 282, 15315-8.
- Meylan, E. and Tschopp, J. (2006) Mol Cell 22, 561-9.
- Thompson, A.J. and Locarnini, S.A. (2007) Immunol Cell Biol 85, 435-45.
- Imaizumi, T. et al. (2002) Biochem Biophys Res Commun 292, 274-9.
- Zhang, X. et al. (2000) Microb Pathog 28, 267-78.
- Yoneyama, M. et al. (2005) J Immunol 175, 2851-8.
- Yoneyama, M. et al. (2004) Nat Immunol 5, 730-7.
- Hornung, V. et al. (2006) Science 314, 994-7.
- Pichlmair, A. et al. (2006) Science 314, 997-1001.
- Kato, H. et al. (2006) Nature 441, 101-5.
- Childs, K. et al. (2007) Virology 359, 190-200.
- Meylan, E. et al. (2005) Nature 437, 1167-72.
- Xu, L.G. et al. (2005) Mol Cell 19, 727-40.
- Kawai, T. et al. (2005) Nat Immunol 6, 981-8.
- Seth, R.B. et al. (2005) Cell 122, 669-82.
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For Research Use Only. Not For Use In Diagnostic Procedures.
Cell Signaling Technology® is a trademark of Cell Signaling Technology, Inc.
Select rabbit monoclonal antibodies are developed, validated, and produced at CST using in part technology under license (granting certain rights including those under U.S. Patents No. 5,675,063 and in some instances 7,429,487) from Epitomics, Inc.