Technical datasheet
Description
Toll-like receptors (TLR) are highly conserved throughout evolution and have been implicated in the innate defense to many pathogens. In Drosophila toll is required for the anti-fungal response, while the related 18-wheeler is involved in antibacterial defenses. In mammals, TLR identified as type I transmembrane signaling receptors with pattern recognition capabilities, have been implicated in the innate host defense to pathogens. TLR4 has been identified next to MD-2 and CD14 as a receptor that is central to the innate immune response to lipopolysaccharides (LPS) of Gram-negative bacteria. The HTA125 monoclonal antibody reacts preferentially, especially in flow cytometry, with human TLR4 (CD284) that is associated with MD-2. HTA125 is a TLR4 (CD284) function-blocking antibody that is useful for studies on the role of TLR4 (CD284) as a receptor for LPS induced cytokine production by TLR4 bearing cells. The antibody was shown to precipitate TLR4 (100 kDa). The antibody HTA125 is cross reactive with canine, cynomolgus monkey, rhesus monkey and marmoset monkey.
Cross Reactivity
| Cross reactant |
Reactivity |
| Canine |
Yes |
| Cynomolgus monkey |
Yes |
| Rhesus monkey |
Yes |
| Marmoset monkey |
Yes |
Immunogen
BALB/c mice were immunized with the Ba/F3 line expressing TLR4
Formulation
0.5 ml (100 µg/ml) 0.2 µm filtered biotinylated antibody solution in PBS, containing 0.02% chlooracetamide and 0.1% bovine serum albumin.
Species
Mouse IgG2a
Application
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F
|
FC1,3,8
|
FS2,3,4,9
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IA5
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IF6
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IP1
|
P
|
W7
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Yes
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No
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N.D.
|
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N.D.= Not Determined; F = Frozen sections; FC = Flow Cytometry; FS = Functional Studies; IA = Immuno Assays; IF = Immuno Fluorescence; IP = Immuno Precipitation; P = Paraffin sections; W = Western blot
Application notes
FC: 300000 cells/50µl were stained with 2 µg antibody for 30 minutes at 4°C
FS: In cell culture 10 µg/ml
IF: Oregon green labeled HTA125 was used in FRAP measurements
IP: HTA125 (4 mg/ml) coupled to Sepharose 4FF beads was added to cell lysate and incubated for 2 hours at 4°C
W: 20 mg protein was analyzed on SDS-PAGE and transferred to nitrocellulose. Blot was blocked with TBS/5% dry milk/0.1% tween-20
Use
For flow cytometry and immunohistology dilutions to be used depend on detection system applied. It is recommended that users test the reagent and determine their own optimal dilutions. The typical starting working dilution is 1:50.
Aliases
TLR4, CD284, ARMD10
Negative control
HEK293 cells
Positive control
Macrophages
Storage and stability
Product should be stored at 4°C. Under recommended storage conditions, product is stable for one year.
References
1. Shimazu, R et al; MD-2, a molecule that confers lipopolysaccharide responsiveness on Toll-like receptor 4. J Exp Med 1999, 189: 1777
2. Tabeta, K et al; Toll-like receptors confer responsiveness to lipopolysaccharide from porphyromonas gingivalis in human gingival fibroblasts. Infect Immun 2000, 68: 3731
3. Akashi, S et al; Regulatory roles for CD14 and phosphatidylinositol in the signaling via Toll-like receptor 4-MD-2. Biochem Biophys Res Commun 2000, 268: 172
4. Wang, J et al; Involvement of CD14 and Toll-like receptors in activation of human monocytes by Aspergillus fumigatus hyphae. Infect Immun 2001, 69: 2402
5. Walton, K et al; Receptors involved in the oxidized 1-palmitoyl-2 arachidonoyl-sn-glycero-3-phosphorylcholine-mediated synthesis of interleukin-8. J Biol Chem 2003, 278: 29661
6. Triantafilou, M et al; Lateral diffusion of Toll-like receptors reveals that they are transiently confined within lipid rafts on the plasma membrane. J Cell Sci 2004, 117: 4007
7. Elner, S et al; TLR4 mediates human retinal pigment epithelial endotoxin binding and cytokine expression. Trans Am Ophthalmol Soc 2005, 103: 126
8. Burgener, I et al; Antibodies specific for human or murine Toll-like receptors detect canine leukocytes by flow cytometry. Vet Immunol Immunopathol 2008, 124: 184
9. Brüll, F et al; TLR2 activation is essential to induce a Th1 shift in human peripheral blood mononuclear cells by plant stanols and plant sterols. J Biol Chem 2010, 285: 2951
Precautions
For research use only. Not for use in or on humans or animals or for diagnostics. It is the responsibility of the user to comply with all local/state and Federal rules in the use of this product. Hycult Biotech is not responsible for any patent infringements that might result with the use of or derivation of this product.
Also available
References
1. Shimazu, R et al; MD-2, a molecule that confers lipopolysaccharide responsiveness on Toll-like receptor 4. J Exp Med 1999, 189: 1777
2. Tabeta, K et al; Toll-like receptors confer responsiveness to lipopolysaccharide from porphyromonas gingivalis in human gingival fibroblasts. Infect Immun 2000, 68: 3731
3. Akashi, S et al; Regulatory roles for CD14 and phosphatidylinositol in the signaling via Toll-like receptor 4-MD-2. Biochem Biophys Res Commun 2000, 268: 172
4. Wang, J et al; Involvement of CD14 and Toll-like receptors in activation of human monocytes by Aspergillus fumigatus hyphae. Infect Immun 2001, 69: 2402
5. Walton, K et al; Receptors involved in the oxidized 1-palmitoyl-2 arachidonoyl-sn-glycero-3-phosphorylcholine-mediated synthesis of interleukin-8. J Biol Chem 2003, 278: 29661
6. Triantafilou, M et al; Lateral diffusion of Toll-like receptors reveals that they are transiently confined within lipid rafts on the plasma membrane. J Cell Sci 2004, 117: 4007
7. Elner, S et al; TLR4 mediates human retinal pigment epithelial endotoxin binding and cytokine expression. Trans Am Ophthalmol Soc 2005, 103: 126
8. Burgener, I et al; Antibodies specific for human or murine Toll-like receptors detect canine leukocytes by flow cytometry. Vet Immunol Immunopathol 2008, 124: 184
9. Brüll, F et al; TLR2 activation is essential to induce a Th1 shift in human peripheral blood mononuclear cells by plant stanols and plant sterols. J Biol Chem 2010, 285: 2951