Description
Human Low-density lipoprotein receptor-related protein 5 (LRP5) ELISA Kit
The Human Low-Density Lipoprotein Receptor Related Protein 5 (LRP5) ELISA Kit is specifically designed for the accurate and reliable detection of LRP5 levels in human serum, plasma, and cell culture supernatants. With its high sensitivity and specificity, this kit provides precise results for a variety of research applications.LRP5 is a key receptor involved in regulating lipid metabolism and bone formation. Dysregulation of LRP5 has been linked to various diseases such as osteoporosis, Alzheimer's disease, and atherosclerosis.
Therefore, measuring LRP5 levels can provide valuable insights into these conditions and aid in the development of potential therapeutic strategies.Overall, the Human LRP5 ELISA Kit is an essential tool for researchers studying lipid metabolism, bone health, and related diseases. Its accuracy and reliability make it a valuable resource for advancing scientific knowledge and developing new treatment options.
| Product Name: | Human Low-density lipoprotein receptor-related protein 5 (LRP5) ELISA Kit |
| SKU: | HUEB2420 |
| Size: | 96T |
| Target: | Human Low-density lipoprotein receptor-related protein 5 (LRP5) |
| Synonyms: | Low-density lipoprotein receptor-related protein 7, LRP-7, LRP-5, LR3, LRP7 |
| Assay Type: | Sandwich |
| Detection Method: | ELISA |
| Reactivity: | Human |
| Detection Range: | 0.312-20ng/mL |
| Sensitivity: | 0.19ng/mL |
| Intra CV: | 4.8% | ||||||||||||||||||||
| Inter CV: | 7.6% | ||||||||||||||||||||
| Linearity: |
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| Recovery: |
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| Function: | Acts as a coreceptor with members of the frizzled family of seven-transmembrane spanning receptors to transduce signal by Wnt proteins (PubMed:11336703, PubMed:11448771, PubMed:15778503, PubMed:11719191, PubMed:15908424, PubMed:16252235). Activates the canonical Wnt signaling pathway that controls cell fate determination and self-renewal during embryonic development and adult tissue regeneration (PubMed:11336703, PubMed:11719191). In particular, may play an important role in the development of the posterior patterning of the epiblast during gastrulation (By similarity). During bone development, regulates osteoblast proliferation and differentiation thus determining bone mass (PubMed:11719191). Mechanistically, the formation of the signaling complex between Wnt ligand, frizzled receptor and LRP5 coreceptor promotes the recruitment of AXIN1 to LRP5, stabilizing beta-catenin/CTNNB1 and activating TCF/LEF-mediated transcriptional programs (PubMed:11336703, PubMed:25920554, PubMed:24706814, PubMed:14731402). Acts as a coreceptor for non-Wnt proteins, such as norrin/NDP. Binding of norrin/NDP to frizzled 4/FZD4-LRP5 receptor complex triggers beta-catenin/CTNNB1-dependent signaling known to be required for retinal vascular development (PubMed:27228167, PubMed:16252235). Plays a role in controlling postnatal vascular regression in retina via macrophage-induced endothelial cell apoptosis (By similarity). |
| Uniprot: | O75197 |
| Sample Type: | Serum, plasma, tissue homogenates, cell culture supernates and other biological fluids |
| Specificity: | Natural and recombinant human Low-density lipoprotein receptor-related protein 5 |
| Sub Unit: | Homodimer; disulfide-linked. Forms phosphorylated oligomer aggregates on Wnt-signaling (By similarity). Component of a Wnt-signaling complex that contains a WNT protein, a FZD protein and LRP5 or LRP6. Interacts with FZD8; the interaction is formed on WNT-binding and signaling (PubMed:11448771). Interacts (via the phosphorylated PPPSP motif domains) with AXIN1; the interaction prevents inhibition of beta-catenin phosphorylation and signaling and is enhanced in the presence of GSK3B and WNT1 or WNT3A (PubMed:11336703, PubMed:14731402). Interacts (via beta-propeller regions 3 and 4) with DKK1; the interaction, enhanced by MESD and/or KREMEN, inhibits beta-catenin signaling by preventing GSK3-mediated phosphorylation of the PPPSP motifs and subsequent, AXIN1 binding (PubMed:11448771, PubMed:15778503, PubMed:19746449). Interacts with MESD; the interaction prevents the formation of LRP5 aggregates, targets LRP5 to the plasma membrane and, when complexed with KREMEN2, increases DKK1 binding (PubMed:17488095, PubMed:19746449, PubMed:15143163). Interacts with CSNK1E (PubMed:16513652). Interacts with SOST; the interaction antagonizes canonical Wnt signaling (PubMed:15778503, PubMed:15908424). Interacts with APCDD1 (PubMed:20393562). Interacts with CAPRIN2 (PubMed:18762581). |
| Research Area: | Cancer |
| Subcellular Location: | Membrane Single-pass type I membrane protein Endoplasmic reticulum Chaperoned to the plasma membrane by MESD. |
| Storage: | Please see kit components below for exact storage details |
| Note: | For research use only |
| UniProt Protein Function: | LRP5: Component of the Wnt-Fzd-LRP5-LRP6 complex that triggers beta-catenin signaling through inducing aggregation of receptor- ligand complexes into ribosome-sized signalsomes. Cell-surface coreceptor of Wnt/beta-catenin signaling, which plays a pivotal role in bone formation. The Wnt-induced Fzd/LRP6 coreceptor complex recruits DVL1 polymers to the plasma membrane which, in turn, recruits the AXIN1/GSK3B-complex to the cell surface promoting the formation of signalsomes and inhibiting AXIN1/GSK3- mediated phosphorylation and destruction of beta-catenin. Appears be required for postnatal control of vascular regression in the eye. Required for posterior patterning of the epiblast during gastrulation. Homodimer; disulfide-linked. Forms phosphorylated oligomer aggregates on Wnt-signaling. Component of a Wnt-signaling complex that contains a WNT protein, a FZD protein and LRP5 or LRP6. Interacts with FZD8; the interaction is formed on WNT-binding and signaling. Interacts (via the phosphorylated PPPSP motif domains) with AXIN1; the interaction prevents inhibition of beta-catenin phosphorylation and signaling and is enhanced in the presence of GSK3B and WNT1 or WNT3A. Interacts (via beta-propeller regions 3 and 4) with DKK1; the interaction, enhanced by MESD and/or KREMEN, inhibits beta-catenin signaling by preventing GSK3-mediated phosphorylation of the PPPSP motifs and subsequent, AXIN1 binding. Interacts with MESD; the interaction prevents the formation of LRP5 aggregates, targets LRP5 to the plasma membrane and, when complexed with KREMEN2, increases DKK1 binding. Interacts with CSNK1E. Interacts with SOST; the interaction antagonizes canonical Wnt signaling. Interacts with APCDD1. Widely expressed, with the highest level of expression in the liver and in aorta. Belongs to the LDLR family. |
| UniProt Protein Details: | Protein type:Membrane protein, integral; Receptor, misc. Chromosomal Location of Human Ortholog: 11q13.4 Cellular Component: plasma membrane; receptor complex Molecular Function:protein binding; toxin transporter activity; Wnt receptor activity; Wnt-protein binding Biological Process: bone marrow development; cholesterol homeostasis; glucose catabolic process; negative regulation of osteoblast differentiation; osteoblast development; positive regulation of cell proliferation; positive regulation of fat cell differentiation; positive regulation of mesenchymal cell proliferation; positive regulation of mitosis; positive regulation of transcription from RNA polymerase II promoter; positive regulation of transcription, DNA-dependent; regulation of blood pressure; retina morphogenesis in camera-type eye; Wnt receptor signaling pathway; Wnt receptor signaling pathway through beta-catenin Disease: Bone Mineral Density Quantitative Trait Locus 1; Endosteal Hyperostosis, Autosomal Dominant; Exudative Vitreoretinopathy 1; Exudative Vitreoretinopathy 4; Osteopetrosis, Autosomal Dominant 1; Osteoporosis; Osteoporosis-pseudoglioma Syndrome; Van Buchem Disease, Type 2 |
| NCBI Summary: | This gene encodes a transmembrane low-density lipoprotein receptor that binds and internalizes ligands in the process of receptor-mediated endocytosis. This protein also acts as a co-receptor with Frizzled protein family members for transducing signals by Wnt proteins and was originally cloned on the basis of its association with type 1 diabetes mellitus in humans. This protein plays a key role in skeletal homeostasis and many bone density related diseases are caused by mutations in this gene. Mutations in this gene also cause familial exudative vitreoretinopathy. Alternative splicing results in multiple transcript variants. [provided by RefSeq, May 2014] |
| UniProt Code: | O75197 |
| NCBI GenInfo Identifier: | 62512139 |
| NCBI Gene ID: | 4041 |
| NCBI Accession: | O75197.2 |
| UniProt Secondary Accession: | O75197,Q96TD6, Q9UES7, Q9UP66, |
| UniProt Related Accession: | O75197 |
| Molecular Weight: | 179,145 Da |
| NCBI Full Name: | Low-density lipoprotein receptor-related protein 5 |
| NCBI Synonym Full Names: | LDL receptor related protein 5 |
| NCBI Official Symbol: | LRP5 |
| NCBI Official Synonym Symbols: | HBM; LR3; OPS; EVR1; EVR4; LRP7; OPPG; BMND1; LRP-5; OPTA1; VBCH2 |
| NCBI Protein Information: | low-density lipoprotein receptor-related protein 5 |
| UniProt Protein Name: | Low-density lipoprotein receptor-related protein 5 |
| Protein Family: | Low-density lipoprotein receptor-related protein |
| UniProt Gene Name: | LRP5 |
| UniProt Entry Name: | LRP5_HUMAN |
| 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. |
| ELISA |
| Human LRP5 ELISA Kit |
