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Detection of TLR4 expression on human peripheral monocytes by purified anti-human TLR4 antibody HTA125 (Cat. No. 551964). Human PBMC were stained with 0.5 µg of purified HTA125 using 3-step staining protocol outlined below and anti-human CD14-FITC (Cat. No. 555397) and analyzed with a BD FACScan™ Flow Cytometer (Becton Dickinson, San Jose, CA).
BD Pharmingen™ Purified Mouse Anti-Human Toll-Like Receptor 4
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准备和存储
推荐的实验流程
A multistep step staining procedure is recommended to amplify immunofluorescent signals for the flow cytometric analysis of human TLR4 expression:
Step 1: Incubate 10^6 cells with 0.25 - 1.0 µg/test of purified HTA125 antibody at 4°C for at least 15 - 20 minutes. Wash cells two times with
staining medium containing sodium azide (e.g., BD Pharmingen™ Stain Buffer, Cat. No. 554656).
Step 2: Incubate the cells with biotinylated anti-mouse Ig (Cat. No. 553999) or biotinylated anti-mouse IgG2a, (Cat. No. 553388) at 4°C for 20
minutes. Wash cells two times.
Step 3: Incubate the cells with ≤0.06 µg of streptavidin-phycoerythrin (Cat. No. 554061) at 4°C for 20 minutes. Wash two times. Resuspend cells
in stain buffer and analyze stained cells by flow cytometry using appropriate specificity and compensation controls.
商品通知
- Since applications vary, each investigator should titrate the reagent to obtain optimal results.
- Caution: Sodium azide yields highly toxic hydrazoic acid under acidic conditions. Dilute azide compounds in running water before discarding to avoid accumulation of potentially explosive deposits in plumbing.
- An isotype control should be used at the same concentration as the antibody of interest.
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
配套商品
The HTA125 monoclonal antibody specifically binds to the human Toll-like receptor 4 (TLR4), also known as CD284. Toll-like receptors (TLR) are type I transmembrane proteins characterized by an extracellular domain containing leucine-rich repeats and a cytoplasmic domain similar to IL-1R family. In humans, ten TLR have been identified so far. The TLR family acts as a pattern-recognition receptor that is an essential component of pathogen recognition and innate immunity. TLR4 has been identified as the receptor for Gram-negative bacterial lipopolysaccharide (LPS). TLR4 mRNA expression has been found in spleen, placenta, ovary, intestine and lung. Using HTA125, TLR4 protein expression has been detected in monocytes, B cells and T cells in human peripheral blood. The human TLR4 gene has been mapped to chromosome 9q32. The immunogen used to generate the HTA125 hybridoma was human TLR4-transfected Ba/F3 cells. The HTA125 antibody reportedly blocks LPS-triggered, TLR4-mediated stimulation of cells.
研发参考 (11)
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Akira S. Toll-like receptors and innate immunity. Adv Immunol. 2001; 78:1-56. (Biology). 查看参考
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Beutler B. Tlr4: central component of the sole mammalian LPS sensor. Curr Opin Immunol. 2000; 12(1):20-26. (Biology). 查看参考
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Chuang T, Ulevitch RJ. Identification of hTLR10: a novel human Toll-like receptor preferentially expressed in immune cells. Biochim Biophys Acta. 2001; 1518(1-2):157-161. (Biology). 查看参考
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Chuang TH, Ulevitch RJ. Cloning and characterization of a sub-family of human toll-like receptors: hTLR7, hTLR8 and hTLR9. Eur Cytokine Netw. 2000; 11(3):372-378. (Biology). 查看参考
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Du X, Poltorak A, Wei Y, Beutler B. Three novel mammalian toll-like receptors: gene structure, expression, and evolution. Eur Cytokine Netw. 2000; 11(3):362-371. (Biology). 查看参考
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Paik YH, Schwabe RF, Bataller R, Russo MP, Jobin C, Brenner DA.. Toll-like receptor 4 mediates inflammatory signaling by bacterial lipopolysaccharide in human hepatic stellate cells.. Hepatology. 2003; 37(5):1043-1055. (Clone-specific: Blocking). 查看参考
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Rock FL, Hardiman G, Timans JC, Kastelein RA, Bazan JF. A family of human receptors structurally related to Drosophila Toll. Proc Natl Acad Sci U S A. 1998; 95(2):588-593. (Biology). 查看参考
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Shimazu R, Akashi S, Ogata H, et al. MD-2, a molecule that confers lipopolysaccharide responsiveness on Toll-like receptor 4. J Exp Med. 1999; 189(11):1777-1782. (Immunogen). 查看参考
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Takeuchi O, Kawai T, Sanjo H, et al. TLR6: A novel member of an expanding toll-like receptor family. Gene. 1999; 231(1-2):59-65. (Biology). 查看参考
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Zarember KA, Godowski PJ. Tissue expression of human Toll-like receptors and differential regulation of Toll-like receptor mRNAs in leukocytes in response to microbes, their products, and cytokines. J Immunol. 2002; 168(2):554-561. (Biology). 查看参考
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Comparisons, where applicable, are made against older BD Technology, manual methods or are general performance claims. Comparisons are not made against non-BD technologies, unless otherwise noted.
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