Skip to main content

Key Product Details

Validated by

Biological Validation

Species Reactivity

Human, Mouse, Rat, Rhesus Macaque

Applications

Immunocytochemistry/ Immunofluorescence, Immunohistochemistry, Immunohistochemistry-Paraffin, Proximity Ligation Assay, Western Blot

Label

Unconjugated

Antibody Source

Polyclonal Rabbit IgG

Concentration

1.0 mg/ml

Product Summary for TLR4 Antibody

Immunogen

This TLR4 antibody was developed against a sythetic peptide corresponding to amino acids in a range between 30-80 of mouse TLR4.

Reactivity Notes

Rat reactivity reported in scientific literature (PMID: 22427516)

Clonality

Polyclonal

Host

Rabbit

Isotype

IgG

Theoretical MW

95.7 kDa.
Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.

Scientific Data Images for TLR4 Antibody

Western Blot: TLR4 Antibody [NB100-56580] - Immunoblot data examining response of primary macrophages from WT and miR-223 knockout mice (KO) to LPS and combined IFNG/LPS; 50ug of whole cell lysate was loaded per lane and run on a denaturing SDS PAGE gel. Densitometry and statistics are presented in (D). All values are expressed as +/- SEM; * indicates p<0.05 via student t-test. Image collected and cropped by CiteAb from the following publication (//doi.org/10.1371/journal.pone.0165962) licensed under a CC-BY license.
Immunohistochemistry-Paraffin: TLR4 Antibody [NB100-56580] - Analysis of TLR4 in mouse kidney tissue using TLR4 antibody at 5 ug/mL.
Western Blot: TLR4 Antibody [NB100-56580] - Analysis of TLR4. (A) 1 ug/lane partial recombinant mouse TLR4 protein (antibody at 2 ug/mL) and (B) RAW cell lysate and (C) Daudi cell lysate (antibody at 5 ug/mL on tumor cell lysate).

Applications for TLR4 Antibody

Application
Recommended Usage

Immunocytochemistry/ Immunofluorescence

2 - 5 ug/mL

Immunohistochemistry-Paraffin

5 ug/mL

Western Blot

2 - 5 ug/mL
Application Notes
This antibody was tested against partial recombinant mouse TLR4 (extracellular portion), and a 75-80 kDa band was observed. Full-length TLR4 is observed at approximately 90 kDa. Zager et al. 2006 (PMID: 16638912) observed TLR4 as an ~95 kDa doublet which is thought to represent different degrees of glycosylation. Presumptive TLR4 30 and 60 kDa cleavage fragments are described in Zagar et al 2007 (PMID: 16885150). Use in PLA reported in scientific literature (PMID:33483465)
Please Note: Optimal dilutions of this antibody should be experimentally determined.

Reviewed Applications

Read 1 review rated 3 using NB100-56580 in the following applications:

Published Applications

Read 25 publications using NB100-56580 in the following applications:

Formulation, Preparation, and Storage

Purification

Immunogen affinity purified

Formulation

PBS

Preservative

0.05% Sodium Azide

Concentration

1.0 mg/ml

Shipping

The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.

Stability & Storage

Store at 4C short term. Aliquot and store at -20C long term. Avoid freeze-thaw cycles.

Background: TLR4

TLR4 (Toll-like receptor 4) is a type-1 transmembrane glycoprotein that is a pattern recognition receptor (PRR) belonging to the TLR family (1-3). TLR4 is expressed in many tissues and is most abundantly expressed in the placenta, spleen, and peripheral blood leukocytes (1). Human TLR4 is synthesized as a 839 amino acid (aa) protein containing a signal sequence (1-23 aa), an extracellular domain (ECD) (24-631 aa), a transmembrane domain (632-652 aa), and Toll/interleukin-1 receptor (TIR) cytoplasmic domain (652-839 aa) with a theoretical molecular weight of 95 kDa (3, 4). The ECD contains 21 leucine-rich repeats (LRRs) and has a horseshoe-shaped structure (3, 4). TLR4 requires binding with the co-receptor myeloid differentiation protein 2 (MD2) largely via hydrophilic interactions for proper ligand sensing and signaling (2-4). In general, the TLR family plays a role in activation of innate immunity and responds to a variety of pathogen-associated molecular patterns (PAMPs) (5). TLR4 is specifically responsive to lipopolysaccharide (LPS), which is found on the outer-membrane of most ram-negative bacteria (3-5). Activation of TLR4 requires binding of a ligand, such as LPS to MD2, followed by MD2-LPS complex binding to TLR4, resulting in a partial complex (TLR4-MD2/LPS) (3, 5). To become fully active, two partial complexes must dimerize thereby allowing the TIR domains of TLR4 to bind other adapter molecular and initiate signaling, triggering an inflammatory response and cytokine production (3, 5).

TLR4 signaling occurs through two distinct pathways: The MyD88 (myeloid differentiation primary response gene 88)-dependent pathway and the MyD88-independent (TRIF-dependent, TIR domain-containing adaptor inducing IFN-beta) pathway (3, 5-7). The MyD88-dependent pathway occurs mainly at the plasma membrane and involves the binding of MyD88-adaptor-like (MAL) protein followed by a signaling cascade that results in the activation of transcription factors including nuclear factor-kappaB (NF-kappaB) that promote the secretion of inflammatory molecules and increased phagocytosis (5-7). Conversely, the MyD88-independent pathway occurs after TLR4-MD2 complex internalization in the endosomal compartment. This pathway involves the binding of adapter proteins TRIF and TRIF-related adaptor molecule (TRAM), a signaling activation cascade resulting in IFN regulatory factor 3 (IRF3) translocation into the nucleus, and secretion of interferon-beta (INF-beta) genes and increased phagocytosis (5-7).

Given its expression on immune-related cells and its role in inflammation, TLR4 activation can contribute to various diseases (6-8). For instance, several studies have found that TLR4 activation is associated with neurodegeneration and several central nervous system (CNS) pathologies, including Alzheimer's disease, Parkinson's disease, and Huntington's disease (6, 7). Furthermore, TLR4 mutations have been shown to lead to higher rates of infections and increased susceptibility to sepsis (7-8). One potential therapeutic approach aimed at targeting TLR4 and neuroinflammation is polyphenolic compounds which include flavonoids and phenolic acids and alcohols (8).

Alternative names for TLR4 includes 76B357.1, ARMD10, CD284 antigen, CD284, EC 3.2.2.6, homolog of Drosophila toll, hToll, toll like receptor 4 protein, TOLL, toll-like receptor 4.

References

1. Vaure, C., & Liu, Y. (2014). A comparative review of toll-like receptor 4 expression and functionality in different animal species. Frontiers in immunology. https://doi.org/10.3389/fimmu.2014.00316

2. Park, B. S., & Lee, J. O. (2013). Recognition of lipopolysaccharide pattern by TLR4 complexes. Experimental & molecular medicine. https://doi.org/10.1038/emm.2013.97

3. Krishnan, J., Anwar, M.A., & Choi, S. (2016) TLR4 (Toll-Like Receptor 4). In: Choi S. (eds) Encyclopedia of Signaling Molecules. Springer, New York, NY. https://doi.org/10.1007/978-1-4614-6438-9_592-1

4. Botos, I., Segal, D. M., & Davies, D. R. (2011). The structural biology of Toll-like receptors. Structure. https://doi.org/10.1016/j.str.2011.02.004

5. Lu, Y. C., Yeh, W. C., & Ohashi, P. S. (2008). LPS/TLR4 signal transduction pathway. Cytokine. https://doi.org/10.1016/j.cyto.2008.01.006

6. Leitner, G. R., Wenzel, T. J., Marshall, N., Gates, E. J., & Klegeris, A. (2019). Targeting toll-like receptor 4 to modulate neuroinflammation in central nervous system disorders. Expert opinion on therapeutic targets. https://doi.org/10.1080/14728222.2019.1676416

7. Molteni, M., Gemma, S., & Rossetti, C. (2016). The Role of Toll-Like Receptor 4 in Infectious and Noninfectious Inflammation. Mediators of inflammation. https://doi.org/10.1155/2016/6978936

8. Rahimifard, M., Maqbool, F., Moeini-Nodeh, S., Niaz, K., Abdollahi, M., Braidy, N., Nabavi, S. M., & Nabavi, S. F. (2017). Targeting the TLR4 signaling pathway by polyphenols: A novel therapeutic strategy for neuroinflammation. Ageing research reviews. https://doi.org/10.1016/j.arr.2017.02.004

Long Name

Toll-like Receptor 4

Alternate Names

CD284

Entrez Gene IDs

7099 (Human); 21898 (Mouse)

Gene Symbol

TLR4

UniProt

Product Documents for TLR4 Antibody

Certificate of Analysis

To download a Certificate of Analysis, please enter a lot number in the search box below.

Product Specific Notices for TLR4 Antibody

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

Loading...
Loading...

Protocols

View specific protocols for TLR4 Antibody (NB100-56580):

Immunocytochemistry Protocol

Culture cells to appropriate density in 35 mm culture dishes or 6-well plates.

1. Remove culture medium and wash the cells briefly in PBS. Add 10% formalin to the dish and fix at room temperature for 10 minutes.
2. Remove the formalin and wash the cells in PBS.
3. Permeablize the cells with 0.1% Triton X100 or other suitable detergent for 10 min.
4. Remove the permeablization buffer and wash three times for 10 minutes each in PBS. Be sure to not let the specimen dry out.
5. To block nonspecific antibody binding, incubate in 10% normal goat serum from 1 hour to overnight at room temperature.
6. Add primary antibody at appropriate dilution and incubate overnight at 4C.
7. Remove primary antibody and replace with PBS. Wash three times for 10 minutes each.
8. Add secondary antibody at appropriate dilution. Incubate for 1 hour at room temperature.
9. Remove secondary antibody and replace with PBS. Wash three times for 10 minutes each.
10. Counter stain DNA with DAPi if required.

FAQs for TLR4 Antibody

Showing  1 - 2 of 2 FAQs Showing All
  • Q: I would like to use this antibody but it has not been validated in my species of interest. Is there any way I can find out if it will work?

    A: We offer risk-free testing of all of our primary antibodies. Please check out our Innovator's Reward Program and test this TLR4 antibody in any unvalidated species or application, without the financial risk of failure.

  • Q: How do I choose secondary antibodies to label the same cells when I have two primary antibodies from the same host?

    A: Use isotype-specific secondary antibodies if the primary antibodies are of different isotypes. You can also make direct conjugates of the primary antibodies by use of antibody labeling kits, dyes, or custom conjugations (please contact Technical Support for custom orders).

Showing  1 - 2 of 2 FAQs Showing All
Loading...