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).

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.

TLR4 Antibody (HTA125) - BSA Free

Novus Biologicals | Catalog # NB100-56723

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Citations (59)

Product Documents

Clone HTA125 was used by HLDA to establish CD designation.

Discontinued Product

NB100-56723 has been discontinued. View all TLR4 products.

Product Specifications
Scientific Data
Applications
Flow Cytometry
Preparation & Storage
Background
Product Documents
Specific Notices
Research Areas
Related Blogs

Key Product Details

Species Reactivity

Validated:

Human, Mouse, Canine

Cited:

Human, Mouse, Rat, Canine

Applications

Validated:

Immunohistochemistry, Immunohistochemistry-Paraffin, Immunohistochemistry Whole-Mount, Western Blot, Block/Neutralize, Flow Cytometry, Flow (Cell Surface), Immunocytochemistry/ Immunofluorescence, Immunoprecipitation, Functional, In vitro assay, CyTOF-ready

Cited:

Immunohistochemistry-Paraffin, Western Blot, ELISA, Block/Neutralize, Flow Cytometry, Flow (Cell Surface), Immunocytochemistry/ Immunofluorescence, Immunoprecipitation, In vitro assay, Functional Assay

Label

Unconjugated

Antibody Source

Monoclonal Mouse IgG_2A Clone # HTA125

Format

BSA Free

View all TLR4 Primary Antibodies »

Product Specifications

Immunogen

This TLR4 Antibody (HTA125) was developed by immunizing mice with Ba/F3 cell line expressing human TLR4 cell surface antigen.

Reactivity Notes

Mouse reactivity reported in scientific literature (PMID: 28219705).

Clonality

Monoclonal

Host

Mouse

Isotype

IgG_2A

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 (HTA125) - BSA Free

Western Blot: TLR4 Antibody (HTA125)BSA Free [NB100-56723]

TLR4-Antibody-HTA125-Western-Blot-NB100-56723-img0023.jpg

TLR4-Antibody-HTA125-Block-Neutralize-NB100-56723-img0022.jpg

Flow Cytometry: TLR4 Antibody (HTA125) - BSA Free [NB100-56723]

Flow Cytometry: TLR4 Antibody (HTA125) [NB100-56723] - Analysis using Azide/BSA FREE version of NB100-56727. Cell surface analysis of TLR4 in stable HEK293/hTLR4 cells using this antibody. Secondary antibody goat anti-mouse IgG-PE at 0.25ug/10^6 cells. Both stable HEK293/hTLR4 and HEK293/Vector cell lines were equally stained.

Flow Cytometry: TLR4 Antibody (HTA125) - BSA Free [NB100-56723]

Flow Cytometry: TLR4 Antibody (HTA125) [NB100-56723] - Staining of U937 cells with mouse anti human CD284:RPE. Image using the PE form of this antibody.

Flow Cytometry: TLR4 Antibody (HTA125) - BSA Free [NB100-56723]

Flow Cytometry: TLR4 Antibody (HTA125) [NB100-56723] - Analysis using Azide/BSA FREE version of NB100-56727. Staining of human peripheral blood monocytes with Mouse anti Human CD284.

Block/Neutralize: TLR4 Antibody (HTA125) [NB100-56723] - Validation of the neutralizing effect of this antibody on TLR4 activity. The TLR4/MD-2/CD14/IL-8 Prom/LUCPorter HeLa cells were plated in 96-well white plates at 1 x 10^5 cells/well. After 16 h, cells were preincubated with different amounts of this antibody for 1 h and then stimulated with 10 ng/ml LPS for 6 h. Luciferase activity was then directly analyzed using the one-step luciferase reporter assay reagent. Data Summary: It inhibited LPS-mediated TLR4 activity in the TLR4/MD-2/CD14/IL-8 Prom/LUCPorter cell line. Image using the Azide Free format of this antibody.

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Applications for TLR4 Antibody (HTA125) - BSA Free

Application

Recommended Usage

Flow (Cell Surface)

reported in scientific literature (PMID 17015691)

Flow Cytometry

0.25 ug / 10^6 cells

Functional

reported in scientific literature (PMID 25412776)

Immunocytochemistry/ Immunofluorescence

reported in scientific literature (PMID 27744428)

Immunohistochemistry Whole-Mount

reported in scientific literature (PMID 26610398)

Immunohistochemistry-Paraffin

reported in scientific literature (PMID 24529879)

Immunoprecipitation

1:10-1:500

In vitro assay

reported in scientific literature (PMID 24676500)

Western Blot

1 - 2 ug/ml. Use reported in scientific literature (PMID 28546218)

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Advanced Features

Spectra Viewer - Custom analysis of spectra from multiple fluorochromes
Spillover Popups - Visualize the spectra of individual fluorochromes
Antigen Density Selector - Match fluorochrome brightness with antigen density

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Formulation, Preparation, and Storage

Purification

Protein G purified

Formulation

PBS

Format

BSA Free

Preservative

0.02% 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, hta125, hta125 tlr4

Gene Symbol

TLR4

Additional TLR4 Products

Product Documents for TLR4 Antibody (HTA125) - BSA Free

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Certificate of Analysis

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Product Specific Notices for TLR4 Antibody (HTA125) - BSA Free

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.

Related Research Areas

Common Macrophage Markers

Diabetes

IL-1 Family

Infectious Diseases

Inflammatory Mediators

NFkB Pathway

NOD-like Receptors and the Inflammasome

Obesity

Pattern Recognition Receptors

Regulatory T Cells (Tregs)

T Cell Antigen Recognition

Toll-Like Receptors

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Protocols

Find general support by application which include: protocols, troubleshooting, illustrated assays, videos and webinars.

FAQs for TLR4 Antibody (HTA125) - BSA Free

Showing 1 - 2 of 2 FAQs Showing All

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).
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).
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.