Description
Mouse Toll-like receptor 1 (Tlr1) ELISA Kit
The Mouse TLR1 (Toll-like Receptor 1) ELISA Kit is specially designed for the precise quantification of TLR1 levels in mouse serum, plasma, and cell culture supernatants. With its outstanding sensitivity and specificity, this kit delivers reliable and reproducible results, making it a valuable tool for a wide range of research purposes.TLR1 is a key protein involved in the immune response, recognizing pathogen-associated molecular patterns and activating the innate immune system.
Its dysregulation has been linked to various infectious and inflammatory diseases, highlighting its importance as a biomarker for studying these conditions and developing potential therapeutic interventions.Overall, the Mouse TLR1 ELISA Kit offers researchers a powerful tool to explore the role of TLR1 in the immune response and its potential as a therapeutic target in various disease settings.
| Product Name: | Mouse Toll-like receptor 1 (Tlr1) ELISA Kit |
| SKU: | MOEB1835 |
| Size: | 96T |
| Target: | Mouse Toll-like receptor 1 (Tlr1) |
| Synonyms: | Toll/interleukin-1 receptor-like protein, TIL, CD281 |
| Assay Type: | Sandwich |
| Detection Method: | ELISA |
| Reactivity: | Mouse |
| Detection Range: | 0.156-10ng/mL |
| Sensitivity: | 0.079ng/mL |
| Intra CV: | 5.3% | ||||||||||||||||||||
| Inter CV: | 10.7% | ||||||||||||||||||||
| Linearity: |
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| Recovery: |
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| Function: | Participates in the innate immune response to microbial agents. Specifically recognizes diacylated and triacylated lipopeptides. Cooperates with TLR2 to mediate the innate immune response to bacterial lipoproteins or lipopeptides. Forms the activation cluster TLR2:TLR1:CD14 in response to triacylated lipopeptides, this cluster triggers signaling from the cell surface and subsequently is targeted to the Golgi in a lipid-raft dependent pathway. Acts via MYD88 and TRAF6, leading to NF-kappa-B activation, cytokine secretion and the inflammatory response (By similarity). Acts as a coreceptor for M.tuberculosis lipoproteins LprG, LpqH and PhoS1 (pstS1), in conjunction with TLR2 and for some but not all lipoproteins CD14 and/or CD36. The lipoproteins act as agonists to modulate antigen presenting cell functions in response to the pathogen (PubMed:19362712). |
| Uniprot: | Q9EPQ1 |
| Sample Type: | Serum, plasma, tissue homogenates, cell culture supernates and other biological fluids |
| Specificity: | Natural and recombinant mouse Toll-like receptor 1 |
| Sub Unit: | Interacts (via extracellular domain) with TLR2. TLR2 seems to exist in heterodimers with either TLR1 or TLR6 before stimulation by the ligand. The heterodimers form bigger oligomers in response to their corresponding ligands as well as further heterotypic associations with other receptors such as CD14 and/or CD36 (By similarity). Binds MYD88 (via TIR domain). Interacts with CNPY3 (PubMed:17998391). |
| Research Area: | Immunology |
| Subcellular Location: | Cell membrane Single-pass type I membrane protein Cytoplasmic vesicle Phagosome membrane Single-pass type I membrane protein Membrane raft Golgi apparatus Does not reside in lipid rafts before stimulation but accumulates increasingly in the raft upon the presence of the microbial ligand. In response to triacylated lipoproteins, TLR2:TLR1 heterodimers are recruited in lipid rafts, this recruitment determine the intracellular targeting to the Golgi apparatus. |
| Storage: | Please see kit components below for exact storage details |
| Note: | For research use only |
| UniProt Protein Function: | TLR1: Participates in the innate immune response to microbial agents. Cooperates with TLR2 to mediate the innate immune response to bacterial lipoproteins or lipopeptides. Acts via MYD88 and TRAF6, leading to NF-kappa-B activation, cytokine secretion and the inflammatory response. Binds MYD88 (via TIR domain). Interacts with CNPY3. Interacts (via extracellular domain) with TLR2. Ligand binding induces the formation of a heterodimer with TLR2. Ubiquitous. Highly expressed in spleen, ovary, peripheral blood leukocytes, thymus and small intestine. Belongs to the Toll-like receptor family. |
| UniProt Protein Details: | Protein type:Membrane protein, integral Cellular Component: Golgi apparatus; integral to plasma membrane; lipid raft Molecular Function:protein heterodimerization activity; transmembrane receptor activity Biological Process: defense response; defense response to bacterium; detection of triacylated bacterial lipoprotein; microglial cell activation; positive regulation of interleukin-6 biosynthetic process; positive regulation of tumor necrosis factor biosynthetic process; response to bacterial lipoprotein; toll-like receptor signaling pathway |
| UniProt Code: | Q9EPQ1 |
| NCBI GenInfo Identifier: | 20140791 |
| NCBI Gene ID: | 21897 |
| NCBI Accession: | Q9EPQ1.1 |
| UniProt Secondary Accession: | Q9EPQ1,Q9EPW5, |
| UniProt Related Accession: | Q9EPQ1 |
| Molecular Weight: | 90,673 Da |
| NCBI Full Name: | Toll-like receptor 1 |
| NCBI Synonym Full Names: | toll-like receptor 1 |
| NCBI Official Symbol: | Tlr1 |
| NCBI Protein Information: | toll-like receptor 1 |
| UniProt Protein Name: | Toll-like receptor 1 |
| UniProt Synonym Protein Names: | Toll/interleukin-1 receptor-like protein; TIL; CD_antigen: CD281 |
| Protein Family: | Toll-like receptor |
| UniProt Gene Name: | Tlr1 |
| UniProt Entry Name: | TLR1_MOUSE |
| Component | Quantity (96 Assays) | Storage |
| ELISA Microplate (Dismountable) | 8×12 strips | -20°C |
| Lyophilized Standard | 2 | -20°C |
| Sample Diluent | 20ml | -20°C |
| Assay Diluent A | 10mL | -20°C |
| Assay Diluent B | 10mL | -20°C |
| Detection Reagent A | 120µL | -20°C |
| Detection Reagent B | 120µL | -20°C |
| Wash Buffer | 30mL | 4°C |
| Substrate | 10mL | 4°C |
| Stop Solution | 10mL | 4°C |
| Plate Sealer | 5 | - |
Other materials and equipment required:
- Microplate reader with 450 nm wavelength filter
- Multichannel Pipette, Pipette, microcentrifuge tubes and disposable pipette tips
- Incubator
- Deionized or distilled water
- Absorbent paper
- Buffer resevoir
*Note: The below protocol is a sample protocol. Protocols are specific to each batch/lot. For the correct instructions please follow the protocol included in your kit.
Allow all reagents to reach room temperature (Please do not dissolve the reagents at 37°C directly). All the reagents should be mixed thoroughly by gently swirling before pipetting. Avoid foaming. Keep appropriate numbers of strips for 1 experiment and remove extra strips from microtiter plate. Removed strips should be resealed and stored at -20°C until the kits expiry date. Prepare all reagents, working standards and samples as directed in the previous sections. Please predict the concentration before assaying. If values for these are not within the range of the standard curve, users must determine the optimal sample dilutions for their experiments. We recommend running all samples in duplicate.
| Step | |
| 1. | Add Sample: Add 100µL of Standard, Blank, or Sample per well. The blank well is added with Sample diluent. Solutions are added to the bottom of micro ELISA plate well, avoid inside wall touching and foaming as possible. Mix it gently. Cover the plate with sealer we provided. Incubate for 120 minutes at 37°C. |
| 2. | Remove the liquid from each well, don't wash. Add 100µL of Detection Reagent A working solution to each well. Cover with the Plate sealer. Gently tap the plate to ensure thorough mixing. Incubate for 1 hour at 37°C. Note: if Detection Reagent A appears cloudy warm to room temperature until solution is uniform. |
| 3. | Aspirate each well and wash, repeating the process three times. Wash by filling each well with Wash Buffer (approximately 400µL) (a squirt bottle, multi-channel pipette,manifold dispenser or automated washer are needed). Complete removal of liquid at each step is essential. After the last wash, completely remove remaining Wash Buffer by aspirating or decanting. Invert the plate and pat it against thick clean absorbent paper. |
| 4. | Add 100µL of Detection Reagent B working solution to each well. Cover with the Plate sealer. Incubate for 60 minutes at 37°C. |
| 5. | Repeat the wash process for five times as conducted in step 3. |
| 6. | Add 90µL of Substrate Solution to each well. Cover with a new Plate sealer and incubate for 10-20 minutes at 37°C. Protect the plate from light. The reaction time can be shortened or extended according to the actual color change, but this should not exceed more than 30 minutes. When apparent gradient appears in standard wells, user should terminatethe reaction. |
| 7. | Add 50µL of Stop Solution to each well. If color change does not appear uniform, gently tap the plate to ensure thorough mixing. |
| 8. | Determine the optical density (OD value) of each well at once, using a micro-plate reader set to 450 nm. User should open the micro-plate reader in advance, preheat the instrument, and set the testing parameters. |
| 9. | After experiment, store all reagents according to the specified storage temperature respectively until their expiry. |
When carrying out an ELISA assay it is important to prepare your samples in order to achieve the best possible results. Below we have a list of procedures for the preparation of samples for different sample types.
| Sample Type | Protocol |
| Serum | If using serum separator tubes, allow samples to clot for 30 minutes at room temperature. Centrifuge for 10 minutes at 1,000x g. Collect the serum fraction and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles. If serum separator tubes are not being used, allow samples to clot overnight at 2-8°C. Centrifuge for 10 minutes at 1,000x g. Remove serum and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles. |
| Plasma | Collect plasma using EDTA or heparin as an anticoagulant. Centrifuge samples at 4°C for 15 mins at 1000 × g within 30 mins of collection. Collect the plasma fraction and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles. Note: Over haemolysed samples are not suitable for use with this kit. |
| Urine & Cerebrospinal Fluid | Collect the urine (mid-stream) in a sterile container, centrifuge for 20 mins at 2000-3000 rpm. Remove supernatant and assay immediately. If any precipitation is detected, repeat the centrifugation step. A similar protocol can be used for cerebrospinal fluid. |
| Cell culture supernatant | Collect the cell culture media by pipette, followed by centrifugation at 4°C for 20 mins at 1500 rpm. Collect the clear supernatant and assay immediately. |
| Cell lysates | Solubilize cells in lysis buffer and allow to sit on ice for 30 minutes. Centrifuge tubes at 14,000 x g for 5 minutes to remove insoluble material. Aliquot the supernatant into a new tube and discard the remaining whole cell extract. Quantify total protein concentration using a total protein assay. Assay immediately or aliquot and store at ≤ -20 °C. |
| Tissue homogenates | The preparation of tissue homogenates will vary depending upon tissue type. Rinse tissue with 1X PBS to remove excess blood & homogenize in 20ml of 1X PBS (including protease inhibitors) and store overnight at ≤ -20°C. Two freeze-thaw cycles are required to break the cell membranes. To further disrupt the cell membranes you can sonicate the samples. Centrifuge homogenates for 5 mins at 5000xg. Remove the supernatant and assay immediately or aliquot and store at -20°C or -80°C. |
| Tissue lysates | Rinse tissue with PBS, cut into 1-2 mm pieces, and homogenize with a tissue homogenizer in PBS. Add an equal volume of RIPA buffer containing protease inhibitors and lyse tissues at room temperature for 30 minutes with gentle agitation. Centrifuge to remove debris. Quantify total protein concentration using a total protein assay. Assay immediately or aliquot and store at ≤ -20 °C. |
| Breast Milk | Collect milk samples and centrifuge at 10,000 x g for 60 min at 4°C. Aliquot the supernatant and assay. For long term use, store samples at -80°C. Minimize freeze/thaw cycles. |
