TLR5 (Human) ELISA Kit (DEIA4322)

Regulatory status: For research use only, not for use in diagnostic procedures.

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Size
96T
Sample
biological fluids, plasma, serum, tissues homogenate
Species Reactivity
Human
Intended Use
This immunoassay kit allows for the in vitro quantitative determination of human Toll-like receptor 5, TLR5 concentrations in serum, plasma, tissue homogenates and other biological fluids.
Contents of Kit
1. Assay plate
2. Standard
3. Sample Diluent
4. Assay Diluent A
5. Assay Diluent B
6. Detection Reagent A
7. Detection Reagent B
8. Wash Buffer (25X)
9. Substrate
10. Stop Solution
11. Plate sealer for 96 wells
12. Instruction
Storage
Store component of the kit at 2-8°C or -20°C upon arrival up to the expiration date. For more detailed information, please download the following document on our website.
Precision
Intra-assay CV: <3.3%
Inter-assay CV: <6.5%
Detection Range
0.312-20 ng/mL
Sensitivity
0.156 ng/mL

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References


Human Toll-like Receptor 8 (TLR8) Is an Important Sensor of Pyogenic Bacteria, and Is Attenuated by Cell Surface TLR Signaling

FRONTIERS IN IMMUNOLOGY

Authors: Moen, Siv H.; Ehrnstrom, Birgitta; Kojen, June F.; Yurchenko, Mariia; Beckwith, Kai S.; Afset, Jan E.; Damas, Jan K.; Hu, Zhenyi; Yin, Hang; Espevik, Terje; Stenvik, Jorgen

TLR8 is an endosomal sensor of RNA degradation products in human phagocytes, and is involved in the recognition of viral and bacterial pathogens. We previously showed that in human primary monocytes and monocyte derived macrophages, TLR8 senses entire Staphylococcus aureus and Streptococcus agalactiae (group B streptococcus, GBS), resulting in the activation of IRF5 and production of IFN beta, IL-12p70, and TNF. However, the quantitative and qualitative impact of TLR8 for the sensing of bacteria have remained unclear because selective inhibitors have been unavailable. Moreover, while we have shown that TLR2 activation attenuates TLR8-IRF5 signaling, the molecular mechanism of this crosstalk is unknown. We here used a recently developed chemical antagonist of TLR8 to determine its role in human primary monocytes challenged with S. aureus, GBS, Streptococcus pneumonia, Pseudomonas aeruginosa, and E. coli. The inhibitor completely blocked cytokine production in monocytes stimulated with TLR8-agonists, but not TLR2-, and TLR4-agonists. Upon challenge with S. aureus, GBS, and S. pneumonia, the TLR8 inhibitor almost eliminated the production of IL-1 beta and IL-12p70, and it strongly reduced the release of IL-6, TNF, and IL-10. With P. aeruginosa infection, the TLR8 inhibitor impaired the production of IL-12p70 and IL-1 beta, while with E. coli infection the inhibitor had less effect that varied depending on the strain and conditions. Signaling via TLR2, TLR4, or TLR5, but not TLR8, rapidly eliminated IRAK-1 detection by immunoblotting due to IRAK-1 modifications during activation. Silencing of IRAK-1 reduced the induction of IFN beta and TNF by TLR8 activation, suggesting that IRAK-1 is required for TLR8-IRF5 signaling. The TLR-induced modifications of IRAK-1 also correlated closely with attenuation of TLR8-IRF5 activation, suggesting that sequestration and/or modification of Myddosome components by cell surface TLRs limit the function of TLR8. Accordingly, inhibition of CD14- and TLR4-activation during E. coli challenge increased the activation of IRF5 and the production of IL-1 beta and IL-12p70. We conclude that TLR8 is a dominating sensor of several species of pyogenic bacteria in human monocytes, while some bacteria attenuate TLR8-signaling via cell surface TLR-activation. Taken together, TLR8 appears as a more important sensor in the antibacterial defense system than previously known.

Single-nucleotide polymorphism-based genetic diversity analysis of the Kilakarsal and Vembur sheep breeds

VETERINARY WORLD

Authors: Selvam, Rathinasamy; Murali, Nagarajan; Thiruvenkadan, A. Kannan; Saravanakumar, Ramesh; Ponnudurai, Gurusamy; Jawahar, Thilak Pon

Aim: The present study was thus undertaken to analyze the genetic diversity of Kilakarsal and Vembur sheep breeds using single-nucleotide polymorphism (SNP) markers within Toll-like receptor (TLR) 3, 5, 6, 9, and 10 genes. Materials and Methods: Competitive allele-specific polymerase chain reaction (PCR)-based end-point genotyping was performed using real-time PCR to type the SNPs. Allele discrimination module implemented in real-time PCR was utilized to call the genotypes based on fluorescence intensity recorded for each of the two alleles. Basic diversity indices, namely, gene frequencies, observed heterozygosity, expected heterozygosity, and inbreeding coefficient (F-IS), and testing for Hardy-Weinberg equilibrium (HWE) were estimated using package for elementary analysis of SNP data software program. Results: Of the 25 SNPs, 22 were found to be polymorphic, whereas two SNPs, namely, TLR3_1081_AC and TLR9_2036_CT, were monomorphic in both Kilakarsal and Vembur sheep populations. The SNP TLR10_1180_AG was monomorphic in Kilakarsal but polymorphic in Vembur sheep. The observed heterozygosities were estimated as 0.289 and 0.309 in Kilakarsal and Vembur sheep, respectively, whereas the expected heterozygosity values were 0.305 and 0.309 in the two breeds, respectively. The overall mean F-IS was 0.107 ranging from -0.005 to 0.241 in Kilakarsal sheep and -0.047 ranging from -0.005 to 0.255 in Vembur sheep. In Kilakarsal sheep, the test for HWE revealed TLR9_1308_GC SNP locus with significant deviation (p< 0.05) due to heterozygosity deficit. In Vembur sheep, TLR10_82_CT and TLR10_292_CG loci showed significant deviation (p< 0.05) due to heterozygosity excess. Other SNP loci did not deviate from HWE (p> 0.05) revealing that the population was in HWE proportions. Conclusions: The SNP markers within five TLR genes (TLR3, TLR5, TLR6, TLR9, and TLR10) utilized for genotyping in this study were highly polymorphic in Kilakarsal and Vembur breeds of sheep. This study on the genetic diversity analysis of the Kilakarsal and Vembur sheep breeds revealed considerable genetic variation within the breeds and it can be utilized to improve desirable traits.

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