Description
Rat Vascular endothelial growth factor receptor 2 (Kdr) ELISA Kit
The Rat Vascular Endothelial Growth Factor Receptor 2 (KDR) ELISA Kit from Assay Genie is a cutting-edge tool designed for precise measurement of KDR levels in rat samples. With its superior sensitivity and specificity, this kit delivers consistent and accurate results, making it an invaluable asset for a variety of research applications.KDR, also known as VEGFR-2, is a key receptor involved in the regulation of vascular endothelial growth factor (VEGF) signaling pathways. It plays a critical role in angiogenesis, vasculogenesis, and endothelial cell survival, making it a crucial target for studying conditions such as cancer, cardiovascular diseases, and ocular disorders.
By utilizing the Rat Vascular Endothelial Growth Factor Receptor 2 (KDR) ELISA Kit, researchers can gain valuable insights into the role of KDR in various physiological and pathological processes, paving the way for the development of novel therapeutic strategies. Trust Assay Genie for reliable and accurate results in your KDR research.
| Product Name: | Rat Vascular endothelial growth factor receptor 2 (Kdr) ELISA Kit |
| SKU: | RTEB0116 |
| Size: | 96T |
| Target: | Rat Vascular endothelial growth factor receptor 2 (Kdr) |
| Synonyms: | Fetal liver kinase 1, Protein-tyrosine kinase receptor flk-1, FLK-1, CD309, VEGFR-2, Flk1 |
| Assay Type: | Sandwich |
| Detection Method: | ELISA |
| Reactivity: | Rat |
| Detection Range: | 78-5000pg/mL |
| Sensitivity: | 39.1pg/mL |
| Intra CV: | 5.1% | ||||||||||||||||||||
| Inter CV: | 9.8% | ||||||||||||||||||||
| Linearity: |
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| Recovery: |
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| Function: | Tyrosine-protein kinase that acts as a cell-surface receptor for VEGFA, VEGFC and VEGFD. Plays an essential role in the regulation of angiogenesis, vascular development, vascular permeability, and embryonic hematopoiesis. Promotes proliferation, survival, migration and differentiation of endothelial cells. Promotes reorganization of the actin cytoskeleton. Isoforms lacking a transmembrane domain may function as decoy receptors for VEGFA, VEGFC and/or VEGFD. Modulates FLT1 and FLT4 signaling by forming heterodimers. Binding of vascular growth factors to isoform 1 leads to the activation of several signaling cascades. Activation of PLCG1 leads to the production of the cellular signaling molecules diacylglycerol and inositol-1,4,5-trisphosphate and the activation of protein kinase C. Mediates activation of MAPK1/ERK2, MAPK3/ERK1 and the MAP kinase signaling pathway, as well as of the AKT1 signaling pathway. Mediates phosphorylation of PIK3R1, the regulatory subunit of phosphatidylinositol 3-kinase, reorganization of the actin cytoskeleton and activation of PTK2/FAK1. Required for VEGFA-mediated induction of NOS2 and NOS3, leading to the production of the signaling molecule nitric oxide (NO) by endothelial cells. Phosphorylates PLCG1. Promotes phosphorylation of FYN, NCK1, NOS3, PIK3R1, PTK2/FAK1 and SRC. |
| Uniprot: | O08775 |
| Sample Type: | Serum, plasma, tissue homogenates, cell culture supernates and other biological fluids |
| Specificity: | Natural and recombinant rat Vascular endothelial growth factor receptor 2 |
| Sub Unit: | Homodimer in the presence of bound dimeric VEGFA, VEGFC or VEGFD ligands; monomeric in the absence of bound ligands. Can also form heterodimers with FLT1/VEGFR1 and FLT4/VEGFR2. Interacts (tyrosine phosphorylated) with LFYN, NCK1, PLCG1. Interacts (tyrosine-phosphorylated active form preferentially) with DAB2IP (via C2 domain and active form preferentially); the interaction occurs at the late phase of VEGFA response and inhibits KDR/VEGFR2 activity. Interacts with SHBSH2D2A/TSAD, GRB2, MYOF, CBL and PDCD6. Interacts (via C-terminus domain) with ERN1 (via kinase domain); the interaction is facilitated in a XBP1- and vascular endothelial growth factor (VEGF)-dependent manner in endothelial cells. |
| Subcellular Location: | Cell membrane Single-pass type I membrane protein Cytoplasm Nucleus Cytoplasmic vesicle Early endosome Cell junction Endoplasmic reticulum Detected on caveolae-enriched lipid rafts at the cell surface. Is recycled from the plasma membrane to endosomes and back again. Phosphorylation triggered by VEGFA binding promotes internalization and subsequent degradation. VEGFA binding triggers internalization and translocation to the nucleus. Localized with RAP1A at cell-cell junctions. Colocalizes with ERN1 and XBP1 in the endoplasmic reticulum in endothelial cells in a vascular endothelial growth factor (VEGF)-dependent manner (By similarity). |
| Storage: | Please see kit components below for exact storage details |
| Note: | For research use only |
| UniProt Protein Function: | VEGFR2: a receptor tyrosine kinase of the VEGFR family. High affinity receptor for VEGF and VEGF-C. Ligand binding induces autophosphorylation and activation. Activated receptor recruits proteins including Shc, GRB2, PI3K, Nck, SHP-1 and SHP-2. Plays a key role in vascular development and regulation of vascular permeability. |
| UniProt Protein Details: | Protein type:EC 2.7.10.1; Protein kinase, tyrosine (receptor); Kinase, protein; Protein kinase, TK; Membrane protein, integral; TK group; VEGFR family Cellular Component: Golgi apparatus; neuron projection; cell surface; integral to plasma membrane; endoplasmic reticulum; early endosome; cytoplasmic membrane-bound vesicle; lipid raft; cell soma; cytoplasm; plasma membrane; nucleus; cell junction; endosome; external side of plasma membrane Molecular Function:integrin binding; vascular endothelial growth factor receptor activity; protein binding; protein-tyrosine kinase activity; growth factor binding; ATP binding Biological Process: positive regulation of positive chemotaxis; peptidyl-tyrosine phosphorylation; protein amino acid autophosphorylation; cell maturation; positive regulation of calcium-mediated signaling; positive regulation of vasodilation; positive regulation of long-term neuronal synaptic plasticity; calcium ion homeostasis; regulation of cell shape; elevation of cytosolic calcium ion concentration; positive regulation of MAPKKK cascade; positive regulation of focal adhesion formation; ovarian follicle development; positive regulation of cell proliferation; positive regulation of mesenchymal cell proliferation; hemopoiesis; angiogenesis; negative regulation of neuron apoptosis; vasculogenesis; positive regulation of TOR signaling pathway; positive regulation of BMP signaling pathway; aging; response to drug; endothelial cell differentiation; cell migration; positive regulation of neurogenesis; cell fate commitment; embryonic hemopoiesis; male gonad development; regulation of endothelial cell differentiation; positive regulation of phosphoinositide 3-kinase cascade; positive regulation of angiogenesis; branching morphogenesis of a tube; cell migration during sprouting angiogenesis; response to hypoxia; positive regulation of endothelial cell proliferation; positive regulation of protein amino acid phosphorylation; vascular endothelial growth factor receptor signaling pathway; lymph vessel development; surfactant homeostasis; hyaluronan metabolic process; alveolus development; neurite morphogenesis; positive regulation of epithelial cell proliferation; positive regulation of cell migration; negative regulation of apoptosis; lung development; positive regulation of nitric-oxide synthase biosynthetic process |
| NCBI Summary: | receptor for vascular endothelial growth factor (VEGF) [RGD, Feb 2006] |
| UniProt Code: | O08775 |
| NCBI GenInfo Identifier: | 6981128 |
| NCBI Gene ID: | 25589 |
| NCBI Accession: | NP_037194.1 |
| UniProt Related Accession: | O08775 |
| Molecular Weight: | 150,394 Da |
| NCBI Full Name: | vascular endothelial growth factor receptor 2 |
| NCBI Synonym Full Names: | kinase insert domain receptor |
| NCBI Official Symbol: | Kdr |
| NCBI Official Synonym Symbols: | Vegfr-2 |
| NCBI Protein Information: | vascular endothelial growth factor receptor 2; FLK-1; fetal liver kinase 1; kinase insert domain protein receptor; protein-tyrosine kinase receptor flk-1; FLK1 kinase insert domain receptor (VEGF receptor 2); FLK1 kinase insert domain receptor (a type III receptor tyrosine kinase) (VEGF receptor 2) |
| UniProt Protein Name: | Vascular endothelial growth factor receptor 2 |
| UniProt Synonym Protein Names: | Fetal liver kinase 1; FLK-1; Protein-tyrosine kinase receptor flk-1 |
| Protein Family: | Vascular endothelial growth factor receptor |
| UniProt Gene Name: | Kdr |
| UniProt Entry Name: | VGFR2_RAT |
| 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. |
