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Nod-like Receptors (NLRs)
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International Edition

Introduction
The innate immune system comprises several classesof pattern recognition receptors, including Toll-like receptors (TLRs), NOD-like receptors (NLRs), and RIG-1like receptors (RLRs). TLRs recognize microbes on thecell surface and in endosomes, whereas NLRs (alsoknown as Nod-LRRs, NACHT-LRRs, and CATERPILLERs)
and RLRs detect microbial components in the cytosol.
NLRs are characterized by three distinct domains. They
contain a Nod (also known as NACHT) domain and a
LRR domain for ligand recognition. The N-terminal effector domain can be a caspase-recruitment domain(CARD), a pyrin domain (PYD), or a baculoviral inhibitory repeat (BIR)-like domain, all are known to be involved in the regulation of apoptotic and inflammatory signalling.

while Nod2 seems to be a general sensor which isactivated by muramyl dipeptide (MDP), the minimal
motif common to all PGNs of Gram-negative as wellas Gram-positive bacteria. Upon activation, Nod1 andNod2 initiate a pro-inflammatory response throughrecruitment of the receptor-interacting protein 2(RIP-2; RICK, CARDIAK) and subsequent activation of
the NF-κB and MAPK pathways (see Figure). Mutationsin Nod2 have been reported to confer susceptibility
to several chronic inflammatory disorders, including
Crohn’s disease.

Selected Review Articles
Signalling pathways and molecular interactions of NOD1 and NOD2: W.
Strober, et al.; Nat. Rev. Immunol. 6, 9 (2006)  Nod-like proteins in immunity, inflammation and disease: J.H. Fritz, et al.; Nat. Immunol. 7, 1250 (2006)
 Intracellular NOD-like receptors in host defense and disease: T.D. Kanneganti, et al.; Immunity 27, 549 (2007)  Nod1 and Nod2 in innate immunity and human inflammatory disorders: L. Le Bourhis, et al.; Biochem. Soc.
Trans. 35, 1479 (2007)  Mammalian NLR proteins; discriminating foe fromfriend: M. Kaparakis, et al.; Immunol. Cell Biol. 85, 495 (2007)  Intracellular
NOD-like receptors in innate immunity, infection and disease: L. Franchi,
et al.; Cell. Microbiol. 10, 1 (2008)  NLRs at the intersection of cell deathand immunity: J.P. Ting, et al.; Nat. Rev. Immunol. 8, 372 (2008)

Nod1 and Nod2
Nod1 and Nod2 are intracellular PRM (pattern-recognition molecules) of the NLR (Nod-like receptor) family. Both proteins are implicated in the detection ofbacterial peptidoglycan (PGN). Nod1 senses mesodiaminopimelic acid (DAP)-containing peptidoglycan found in the cell wall of Gram-negative bacteria,

Salmonella

highlight

PG

Legionella

Staphylococcus
ToxinsListeria

RNA

ATP Uric acid
Crystals

Flagellin

MDP
DAP

IPAF

NALP3

Nod1
NAIP

Nod2

RIP2

Caspase-1

NF-κB

Caspase-1

MAPK

ProIL-1βCell Death

Inflammatory Response

LRR

Cardinal

Asc

PYD

NACHT

NAD

IL-1β

Inflammatory Response

FIND

CARD

BIR

FIGURE: Activation of cytosolic NLRs by bacterial compounds and “danger”-signals.

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NALPs, NAIP, IPAF & Inflammasomes
Inflammasomes are cytoplasmic multiprotein
complexes that mediate the maturation of theproinflammatory cytokines IL-1β, IL-18 and possibly IL-33 by controlling the activation of theinflammatory caspases-1 and -5. Several inflammasomes have been described and they are defined by the NLR protein that they contain: the
NALP1 inflammasome, the NALP3 inflammasome and the IPAF inflammasome. Upon sensing of their respective ligands, NALP1 and
NALP3 recruit the adaptor protein Asc (apoptosis-associated speck-like protein containing
a CARD) through homophilic PYD-PYD interactions. Asc contains an N-terminal PYD and a Cterminal CARD that allows the recruitment ofinflammatory caspases, especially caspase-1,
to the platform through CARD-CARD interactions. The oligomerization of NALPs is believed
to bring inflammatory caspases into close proximity, leading to their activation within the inflammasome. In contrast to the NALPs, IPAF
does not recruit an adaptor molecule but directly activates caspase-1 via its CARD domain(see Figure). The assembly of the different inflammasomes leads to a common outcome,
namely the activation of caspase-1. Caspase-1
is required for the processing and subsequent
release of active proinflammatory cytokines
such as IL-18 and IL-1β, which is a major mediator of inflammation. IL- 33, a cytokine thatis involved in generating a TH2- cell response,is also cleaved by caspase-1. In addition, inflammasome activation can lead to host cell deathin certain cell types, which might be importantin restricting the intracellular replication of invasive bacterial pathogens.

IPAF
Inflammasome
IPAF

NAIP

NALP1
Inflammasome

NALP3
Inflammasome

NALP1

NALP3
Pyrin
Cardinal
LRR Repeats?

?

BIR Repeats
NACHT Domain
CARD Domain

?
ASC

ASC

PYD Domain
FIIND

B Box, Coiled-coil,
PRY-SPRY Domain

CASP1

CASP1 CASP5

CASP1 CASP1

Caspase Domain

FIGURE: Schematic models of proposed caspase-1-activating inflammasomes. The ligand-sensing motifs (LRR repeats), initiate the formation ofoligomers. PYD-PYD and CARD-CARD homotypic interactions are crucial for the recruitment and activation of either the adaptor Asc or the inflammatory caspases. IPAF and NAIP may be part of the same complex, similarly CARDINAL and Pyrin may be involved in the formation or regulation of
NALP-based inflammasomes. Adapted from Inflammatory caspases and inflammasomes: master switches of inflammation: F. Martinon & J. Tschopp;
Cell Death. Differ. 14, 10 (2007).

Selected Review Articles
Inflammasome adaptors and sensors: intracellular regulators of
infection and inflammation: S. Mariathasan & D.M. Monack; Nat.
Rev. Immunol. 7, 31 (2007)  Inflammatory caspases and inflammasomes: master switches of inflammation: F. Martinon & J. Tschopp;
Cell Death Differ. 14, 10 (2007)  NALP inflammasomes: a central

role in innate immunity: F. Martinon, et al.; Semin. Immunopathol.
29, 213 (2007)  The inflammasome: a danger sensing complex
triggering innate immunity: V. Petrilli, et al.; Curr. Opin. Immunol.
19, 615 (2007)  COPs and POPs: modulators of inflammasome activity: C. Stehlik & A. Dorfleutner; J. Immunol. 179, 7993 (2007)

Nod1 [CARD4] & Nod2 [CARD15]
PAb to Nod1 (human) (AL184)

PAb to Nod2 (human) (NT)PAb to RIP2 (human) (CT)

ALX-210-918-C050
50 μg
From rabbit. IMMUNOGEN: Synthetic peptide corresponding to aa 2-31 (E2EQGHSEMEIIPSESHPHIQLLKSNRELLV31) of human Nod1 (CARD4).
SPECIFICITY: Recognizes human Nod1. APPLICATION: WB.

PSC-2511-C100
100 μgFrom rabbit. IMMUNOGEN: Synthetic peptide corresponding to 16 aa at the N-terminus of human
Nod2 (CARD15). SPECIFICITY: Recognizes human
Nod2. Does not cross-react with Nod1 (CARD4).
APPLICATION: WB. BP: PSC-2511P.

PSC-2183-C100
100 μgFrom rabbit. IMMUNOGEN: Synthetic peptide corresponding to aa 508-522 of C-terminal human
RIP2 (receptor-interacting protein 2). SPECIFICITY: Recognizes human RIP2. Detects a bandof ~60kDa by Western blot. APPLICATION: WB.
BP: PSC-2183P.

RIP2

PAb to RIP2 (NT)MAb to Nod2 (human) (2D9)
ALX-803-307-1
1 Vial
CLONE: 2D9. ISOTYPE: Mouse IgG1. IMMUNOGEN: Recombinant human Nod2 (CARD15) (aa 28-301).
SPECIFICITY: Recognizes human Nod2. APPLICATION: IHC, IP, WB.

PAb to Nod2 (human)
ALX-210-373-1
1 VialFrom rabbit. IMMUNOGEN: Recombinant human
Nod2 (CARD15) (aa 28-301). SPECIFICITY: Recognizes human Nod2. Does not cross-react with
SHAM. APPLICATION: IP, WB.

PAb to Nod2 (human) (CT)
PSC-2513-C100
100 μgFrom rabbit. IMMUNOGEN: Synthetic peptide corresponding to 14 aa at the C-terminus of human
Nod2 (CARD15). SPECIFICITY: Recognizes human
Nod2. Does not cross-react with Nod1 (CARD4).
APPLICATION: WB. BP: PSC-2513P.

2

MAb to RIP2 (human) (Nick-1)
ALX-804-139-C100
100 μg
CLONE: Nick-1. ISOTYPE: Rat IgG2a. IMMUNOGEN: Recombinant human RIP2 (receptor-interacting
protein 2) (aa 1-322). SPECIFICITY: Recognizes
human RIP2. APPLICATION: WB.

PSC-2075-C100
100 μgFrom rabbit. IMMUNOGEN: Synthetic peptide corresponding to aa 11-30 of N-terminal human
RIP2 (receptor-interacting protein 2). SPECIFICITY: Recognizes human, mouse and rat RIP2.
Detects a band of ~60kDa by Western blot.
APPLICATION: WB. BP: PSC-2075P.

LIT: Participation of Rip2 in Lipopolysaccharide signaling is independent of its kinase activity: C. Lu et al.; J. Biol. Chem. 280, 16278 (2005) 
PIDD mediates NF-kB activation in Response to DNA damage: Janssens, et al.; Cell 123, 1079 (2005)

Positive Control (Cell Lysate) for
Antibodies to RIP2 (human)
ALX-840-604-R100
100 μlWhole cell lysate of HEK 293T cells transfected with an expression plasmid for human RIP2
(receptor-interacting protein 2) (aa 1-541).

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Positive Control (Cell Lysate) for
Antibodies to Asc (human)

Antibodies to NALPs
MAb to NALP1 (human) (Nalpy1-4)

PAb to NALP2 (CT)

ALX-804-803-C100
100 μg
CLONE: Nalpy1-4. ISOTYPE: Mouse IgG1. IMMUNOGEN: Recombinant human NALP1 (NACHT-,LRR- and PYD-containing protein 1) (pyrin domain). SPECIFICITY: Recognizes the pyrin domain(PYD) of human NALP1. APPLICATION: IHC (FS,PS), ICC, IP, WB.

PSC-3027-C100
100 μgFrom rabbit. IMMUNOGEN: Synthetic peptide corresponding to 12 aa near the C-terminal of human NALP2 (NACHT-, LRR- and PYD-containing protein 2). SPECIFICITY: Recognizes human,
mouse and rat NALP2. APPLICATION: WB. BP: PSC3027P.

LIT: Inflammasome components NALP 1 and 3 show distinct but separate expression profiles in human tissues suggesting a site-specificrole in the inflammatory response: J.A. Kummer, et al.; J. Histochem.
Cytochem. 55, 443 (2007)

MAb to NALP3 (human) (Nalpy3-a)

PAb to CARD8 (human) (CT)

ALX-804-818-C100
100 μg
CLONE: Nalpy3-a. ISOTYPE: Mouse IgG1. IMMUNOGEN: Recombinant human NALP3 (NACHT-,LRR- and PYD-containing protein 3) (pyrin domain). SPECIFICITY: Recognizes human NALP3.
Detects endogenous protein by ICC and WB.
APPLICATION: ICC, WB.

PSC-3187-C100
100 μgFrom rabbit. IMMUNOGEN: Synthetic peptide corresponding to aa near the C-terminus of human
CARD8 (caspase recruitment domain-containing protein 8). SPECIFICITY: Recognizes human
CARD8. APPLICATION: WB. BP: PSC-3187P.

LIT: NALP3 forms an IL-1beta-processing inflammasome with increased activity in Muckle-Wells autoinflammatory disorder: L. Agostini, et al.; Immunity 20, 319 (2004)

IPAF [CARD12]

ALX-210-904-R100
100 μlFrom rabbit. IMMUNOGEN: Synthetic peptide corresponding to aa 2-25 (A2GGAWGRLACYLEFLKKEELKEFQ25) of N-terminal human NALP1 (NACHT-,LRR- and PYD-containing protein 1). SPECIFICITY:
Recognizes human NALP1. APPLICATION: WB.
LIT: The inflammasome: a molecular platform triggering activationof inflammatory caspases and processing of proIL-beta: F. Martinon;
Mol. Cell. 10, 417 (2002)

PAb to NALP1 (human) (CT) (Bur 242)
ALX-210-017-R050
50 μl
From rabbit. IMMUNOGEN: Synthetic peptide
corresponding to aa 1058-1077 (C1058VPSPASQGDLHTKPLGTDD1077) of C-terminal human NALP1 (NACHT-, LRR- and PYD-containing protein 1). SPECIFICITY: Recognizes human
NALP1. APPLICATION: IHC (PS), IP, WB.

ALX-804-819-C100
100 μg
CLONE: Nalpy3-b. ISOTYPE: Mouse IgG1. IMMUNOGEN: Recombinant human NALP3 (NACHT-,LRR- and PYD-containing protein 3) (pyrin domain). SPECIFICITY: Recognizes human NALP3.
Detects endogenous protein by IP and WB. APPLICATION: IHC (FS), IP, WB.
LIT: NALP3 forms an IL-1beta-processing inflammasome with increased activity in Muckle-Wells autoinflammatory disorder: L.
Agostini, et al.; Immunity 20, 319 (2004)  Inflammasome components NALP 1 and 3 show distinct but separate expression profiles in human tissues suggesting a site-specific role in the inflammatory response: J.A. Kummer, et al.; J. Histochem. Cytochem. 55,
443 (2007)

ALX-210-018-R050
50 μl
From rabbit. IMMUNOGEN: Synthetic peptide
corresponding to aa 161-180 (P161SSPDHESPSQESPNAPTST180) of N-terminal human NALP1(NACHT-, LRR- and PYD-containing protein 1).
SPECIFICITY: Recognizes human NALP1. APPLICATION: IHC (PS), IP, WB.
LIT: A novel enhancer of the Apaf1 apoptosome involved in cytochrome c-dependent caspase activation and apoptosis: Z.L. Chu, etal.; J. Biol. Chem. 276, 9239 (2001)

Positive Control (Cell Lysate) for Antibodies to NALP1 (human)
ALX-840-603-R100
100 μlWhole cell lysate of HEK 293T cells transfectedwith an expression plasmid for human NALP1(NACHT-, LRR- and PYD-containing protein 1)(aa 1-1429).

ALX-804-848-C100
100 μg
CLONE: Luna-1. ISOTYPE: Mouse IgG2b. IMMUNOGEN: Recombinant human IPAF (ice proteaseactivating factor) (aa 9-494). SPECIFICITY: Recognizes human IPAF. APPLICATION: WB.

PAb to IPAF (human)
PSC-3107-C100
100 μgFrom rabbit. IMMUNOGEN: Synthetic peptide corresponding to aa near the C-terminus of human IPAF (ice protease-activating factor). SPECIFICITY: Recognizes human IPAF. APPLICATION:
IHC, WB. BP: PSC-3107P.

NAIP
FIGURE: Staining of endogenous NALP3 in epithelial layer of humantonsil (frozen section)using MAb to NALP3(Nalpy-3b) (Prod. No.
ALX-804-819).

LIT: A novel enhancer of the Apaf1 apoptosome involved in cytochrome c-dependent caspase activation and apoptosis: Z.L. Chu, etal.; J. Biol. Chem. 276, 9239 (2001)

PAb to NALP1 (human) (NT) (Bur 241)

MAb to IPAF (human) (Luna-1)

PAb to NALP12 (human) (AL236)
ALX-210-958-C100
100 μgFrom rabbit. IMMUNOGEN: Synthetic peptide corresponding to aa near the N-terminus of human NALP12. SPECIFICITY: Recognizes human
NALP12. APPLICATION: IP, WB.

Asc
PAb to Asc (AL177)
ALX-210-905-R100
100 μlFrom rabbit. IMMUNOGEN: Synthetic peptide corresponding to aa 2-27 (G2RARDAILDALENLTAEELKKFKLKL27) of N-terminal human Asc (apoptosis-associated speck-likeprotein containing CARD; Pycard). SPECIFICITY: Recognizes human and mouse Asc.
APPLICATION: ICC, IP, WB.
LIT: The inflammasome: a molecular platform triggering activationof inflammatory caspases and processing of proIL-beta: F. Martinon,
et al.; Mol. Cell. 10, 417 (2002)  NALP3 forms an IL-1beta-processing inflammasome with increased activity in Muckle-Wells autoinflammatory disorder: L. Agostini, et al.; Immunity 20, 319 (2004) 
P2X7 Receptor Differentially Couples to Distinct Release Pathwaysfor IL-1{beta} in Mouse Macrophage: P. Pelegrin, et al.; J. Immunol.
180, 7147 (2008)

Purified (PF) = Purified (Preservative free); FC = Flow Cytometry; ICC = Immunocytochemistry; IP = Immunoprecipitation;
IHC = Immunohistochemistry (FS = Frozen Sections, PS = Paraffin Sections); WB = Western blot; BP = Blocking Peptide

PAb to NAIP (human) (CT)PSC-3315-C100
100 μgFrom rabbit. IMMUNOGEN: Synthetic peptide corresponding to 13 aa at the C-terminus of human NAIP (neuronal apoptosis inhibitory protein). SPECIFICITY: Recognizes human NAIP. APPLICATION: WB. BP: PSC-3315P.

Pyrin
PAb to Pyrin (human) (AL196)
ALX-210-946-C100
100 μgFrom rabbit. IMMUNOGEN: Synthetic peptide corresponding to aa 2-29 (A2KTPSDHLLSTLEELVPYDFEKFKFKLQ29) of human pyrin (MEFV; mediterranean fever protein). SPECIFICITY: Recognizes
human pyrin. APPLICATION: IP, WB.

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IL-1

IL-1RII

IL-1RI

sST2?

IL-18bp
IL-18Ab

IL-1RaAnakinra (rIL-Ra)
IL-1 trap (sIL-1R-Fc)

highlight

PAb to NALP1 (human) (AL176)

MAb to NALP3 (human) (Nalpy3-b)

CARD8 [Cardinal]

PRODUC T FLYER

FIGURE: Detection of NALP1 in 293T cells transfected with a human NALP1 expression plasmid. Left: Phase contrast. Right: Staining with MAb to NALP1 (Nalpy1-4).

ALX-840-602-R100
100 μlWhole cell lysate of HEK 293T cells transfected with an expression plasmid for human Asc(apoptosis-associated speck-like protein containing CARD; Pycard) (aa 1-195).

I N N AT E I M M U N I T YInternational Edition

IL-1AcP

TIR
Mimetics

MyD88
IRAK1
IRAK4

ST2

IL-18AcP

IL-18 R

MyD88
IRAK1
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MyD88
IRAK1
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MyD88
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TRAF6

TRAF6

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Proteasomal
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cytokines and theirrole in destructivemmicals.cowww.alexis-bioche3www.alexis-biochemicals.com

Nod-like Receptor Ligands
Nigericin . Na

Ac-muramyl-Ala-D-Glu-amide[Antibiotic K 178]

[MDP-LD; N-Acetylmuramyl-L-alanyl-D-isoglutamine]

ALX-380-050-M005
5 mg
ALX-380-050-M025
25 mg
Isolated from Streptomyces hygroscopicus.

R-848

ALX-151-035-M001
1 mg
ALX-151-035-M005
5 mg
Synthetic. Specific ligand for NALP3. For inactive control compound see MDP-DD (Prod. No.
ALX-151-036).

[4-Amino-2-(ethoxymethyl)-a,a-dimethyl-1H-imidazo[4,5-c]quinoline-1-ethanol; S 28463; Resiquimod]

Adenosine 5‘-triphosphate . 2Na

LIT: The immune response modifier resiquimod mimics CD40-induced B cell
activation: G.A. Bishop, et al.; Cell. Immunol. 208, 9 (2001)  Small anti-viralcompounds activate immune cells via the TLR7 MyD88-dependent signaling
pathway: H. Hemmi, et al.; Nat. Immunol. 3, 196 (2002)  Human TLR7 or TLR8
independently confer responsiveness to the antiviral compound R-848: M. Jurk,
et al.; Nat. Immunol. 3, 499 (2002)  Resiquimod: a new immune response modifier with potential as a vaccine adjuvant for Th1 immune responses: J.J. Wu, etal.; Antiviral Res. 64, 79 (2004)  Direct stimulation of human T cells via TLR5 andTLR7/8: flagellin and R-848 up-regulate proliferation and IFN-gamma production by memory CD4+ T cells: G. Caron, et al.; J. Immunol. 175, 1551 (2005)  TLR
agonists as vaccine adjuvants: comparison of CpG ODN and Resiquimod (R-848):
R.D. Weeratna, et al.; Vaccine 23, 5263 (2005)  Treatment of intravaginal HSV-2
infection in mice: a comparison of CpG oligodeoxynucleotides and resiquimod(R-848): M.J. McCluskie, et al.; Antiviral Res. 69, 77 (2006)[ATP . 2Na]

ALX-480-021-G001
1g
ALX-480-021-G005
5g
Specially crystallized and tested on biological activity.
LIT: Gout-associated uric acid crystals activate the NALP3 inflammasome: F. Martinon, et al.; Nature 440, 237 (2006)

Flagellin (high purity)
ALX-522-058-C010
10 μg
Isolated from Salmonella typhimurium strain 14028.
SPECIFICITY: Binds to human and mouse TLR5. BIOLOGICAL ACTIVITY: Activation of TLR5 in human epithelial cellassays based on NF-κB luciferase fusions.
LIT: Flagellin stimulation of intestinal epithelial cells triggers CCL20-mediatedmigration of dendritic cells: F. Sierro, et al.; PNAS 98, 13722 (2001)  Pathophysiological role of Toll-like receptor 5 engagement by bacterial flagellin in colonicinflammation: S.H. Rhee, et al.; PNAS 102, 13610 (2005)

Imiquimod[R-837]

ALX-420-039-M100
100 mg
ALX-420-039-M250
250 mgTopical immune response modifier that inhibits angiogenesis. Up-regulates IL-18 and down-regulates
MMP-9 through recognition of TLR7 and subsequent
activation of MyD88-dependent pathway.

New NALP3 Inflammasome
Activator

LIT: Imiquimod applied topically: a novel immune response modifier and newclass of drug: R.L. Miller, et al.; Int. J. Immunopharmacol. 21, 1 (1999)  Small
anti-viral compounds activate immune cells via the TLR7 MyD88-dependent
signaling pathway: H. Hemmi, et al.; Nat. Immunol. 3, 196 (2002)  In vivo andin situ modulation of the expression of genes involved in metastasis and angiogenesis in a patient treated with topical imiquimod for melanoma skin metastases: C. Hesling, et al.; Br. J. Dermatol. 150, 761 (2004)  Imiquimod as anantiangiogenic agent: V.W. Li, et al.; J. Drugs Dermatol. 4, 708 (2005)  Imiquimod is a strong inhibitor of tumor cell-induced angiogenesis: S. Majewski, etal.; Int. J. Dermatol. 44, 14 (2005)

ALX-400-047-M002
2 mg
Specially crystallized and tested on biological activity.

Nigericin . Na

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Monosodium urate (crystals)[MSU Crystals; Uric Acid Crystals]

LIT: Gout-associated uric acid crystals activate the NALP3 inflammasome:F. Martinon, et al.; Nature 440, 237 (2006)

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R-848

Imiquimod

Monosodium urate (crystals)
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OCT-08

ALX-420-038-M005
5 mg
ALX-420-038-M025
25 mg
Selective ligand for Toll-like receptor 7 (TLR7) in mouseand for TLR7 and TLR8 in human. Potent antitumor
and antiviral compound. Stimulates antibody secretion, cytokine production, protection from apoptosis
and CD80 upregulation.