Human ERK3 (Extracellular Signal Regulated Kinase 3) ELISA Kit (HUES02998)
- Product type:
- ELISA Kit
- ELISA Type:
|Detection Range:||0.31-20 ng/mL|
|Sample Volume Required Per Well:||100µL|
|Sample Type:||Serum, plasma and other biological fluids|
|Specificity:||This kit recognizes Human ERK3 in samples. No significant cross-reactivity or interference between Human ERK3 and analogues was observed.|
This ELISA kit uses Sandwich-ELISA as the method. The micro ELISA plate provided in this kit has been pre-coated with an antibody specific to Human ERK3. Standards or samples are added to the appropriate micro ELISA plate wells and combined with the specific antibody. Then a biotinylated detection antibody specific for Human ERK3 and Avidin-Horseradish Peroxidase (HRP) conjugate are added to each micro plate well successively and incubated. Free components are washed away. The substrate solution is added to each well. Only those wells that contain Human ERK3, biotinylated detection antibody and Avidin-HRP conjugate will appear blue in color. The enzyme-substrate reaction is terminated by adding Stop Solution and the color turns yellow. The optical density (OD) is measured spectrophotometrically at a wavelength of 450 nm ± 2 nm. The OD value is proportional to the concentration of Human ERK3. The concentration of Human ERK3 in samples can be calculated by comparing the OD of the samples to the standard curve.
|UniProt Protein Function:||ERK1: a serine/threonine kinase of the GMGC group that plays a critical role in the regulation of cell growth and differentiation. ERK1 (MAPK3) and ERK2 (MAPK1) play central roles in MAPK cascades and are activated by a wide variety of extracellular signals including growth and neurotrophic factors, cytokines, hormones and neurotransmitters. Depending on the cellular context, MAPK cascades mediate diverse biological functions such as cell growth, adhesion, survival and differentiation through the regulation of transcription, translation, cytoskeletal rearrangements. MAPK cascades also plays a role in initiation and regulation of meiosis, mitosis, and postmitotic functions in differentiated cells by phosphorylating a number of transcription factors. Activation of MAP kinases occurs through phosphorylation of threonine and tyrosine residues at the sequence T*EY* by upstream MAP kinase kinases, MEK1 and -2. Phosphorylation of both the threonine and tyrosine are required for activity. This phosphorylation causes dramatic conformational changes, which enable full activation and interaction of MAPK1/ERK2 with its substrates.|
|UniProt Protein Details:|
Protein type:EC 2. 7. 11. 24; Protein kinase, CMGC; Protein kinase, Ser/Thr (non-receptor); Kinase, protein; CMGC group; MAPK family; ERK subfamily; MAPK/ERK subfamily
Chromosomal Location of Human Ortholog: 16p11. 2
Cellular Component: microtubule cytoskeleton; nucleoplasm; Golgi apparatus; focal adhesion; cytoskeleton; mitochondrion; late endosome; early endosome; nuclear envelope; caveola; pseudopodium; nucleus; cytosol
Molecular Function:MAP kinase activity; protein binding; phosphotyrosine binding; ATP binding; phosphatase binding
Biological Process: axon guidance; DNA damage induced protein phosphorylation; activation of MAPKK activity; nerve growth factor receptor signaling pathway; positive regulation of histone phosphorylation; viral reproduction; apoptosis; activation of MAPK activity; stress-activated MAPK cascade; toll-like receptor 3 signaling pathway; sensory perception of pain; protein amino acid phosphorylation; toll-like receptor 10 signaling pathway; BMP signaling pathway; toll-like receptor 5 signaling pathway; response to exogenous dsRNA; regulation of transcription factor activity; small GTPase mediated signal transduction; lipopolysaccharide-mediated signaling pathway; toll-like receptor 4 signaling pathway; epidermal growth factor receptor signaling pathway; platelet activation; fibroblast growth factor receptor signaling pathway; cytokine and chemokine mediated signaling pathway; MyD88-independent toll-like receptor signaling pathway; MAPKKK cascade; transcription from RNA polymerase I promoter; cell cycle; toll-like receptor 2 signaling pathway; MyD88-dependent toll-like receptor signaling pathway; regulation of stress-activated MAPK cascade; organ morphogenesis; cartilage development; Ras protein signal transduction; toll-like receptor signaling pathway; insulin receptor signaling pathway; innate immune response; gene expression; positive regulation of histone acetylation; positive regulation of transcription from RNA polymerase II promoter; positive regulation of protein amino acid phosphorylation; toll-like receptor 9 signaling pathway; transcription initiation from RNA polymerase I promoter; vascular endothelial growth factor receptor signaling pathway; blood coagulation; phosphorylation; regulation of cytoskeleton organization and biogenesis
|NCBI Summary:||The protein encoded by this gene is a member of the MAP kinase family. MAP kinases, also known as extracellular signal-regulated kinases (ERKs), act in a signaling cascade that regulates various cellular processes such as proliferation, differentiation, and cell cycle progression in response to a variety of extracellular signals. This kinase is activated by upstream kinases, resulting in its translocation to the nucleus where it phosphorylates nuclear targets. Alternatively spliced transcript variants encoding different protein isoforms have been described. [provided by RefSeq, Jul 2008]|
|NCBI GenInfo Identifier:||232066|
|NCBI Gene ID:||5595|
|NCBI Accession:||P27361. 4|
|UniProt Secondary Accession:||P27361,Q8NHX1, A8CZ58, B0LPG3,|
|UniProt Related Accession:||P27361|
|NCBI Full Name:||Mitogen-activated protein kinase 3|
|NCBI Synonym Full Names:||mitogen-activated protein kinase 3|
|NCBI Official Symbol:||MAPK3|
|NCBI Official Synonym Symbols:||ERK1; ERT2; ERK-1; PRKM3; P44ERK1; P44MAPK; HS44KDAP; HUMKER1A; p44-ERK1; p44-MAPK|
|NCBI Protein Information:||mitogen-activated protein kinase 3; MAPK 1; MAP kinase isoform p44; insulin-stimulated MAP2 kinase; extracellular signal-related kinase 1; extracellular signal-regulated kinase 1; microtubule-associated protein 2 kinase|
|UniProt Protein Name:||Mitogen-activated protein kinase 3|
|UniProt Synonym Protein Names:||ERT2; Extracellular signal-regulated kinase 1; ERK-1; Insulin-stimulated MAP2 kinase; MAP kinase isoform p44; p44-MAPK; Microtubule-associated protein 2 kinase; p44-ERK1|
|UniProt Gene Name:||MAPK3|
|UniProt Entry Name:||MK03_HUMAN|
As the OD values of the standard curve may vary according to the conditions of the actual assay performance (e. g. operator, pipetting technique, washing technique or temperature effects), the operator should establish a standard curve for each test. Typical standard curve and data is provided below for reference only.
Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Human ERK3 were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human ERK3 were tested on 3 different plates, 20 replicates in each plate.
|Intra-assay Precision||Inter-assay Precision|
|C V (%)||5.94||5.93||4.46||4.90||5.26||5.26|
The recovery of Human ERK3 spiked at three different levels in samples throughout the range of the assay was evaluated in various matrices.
|Sample Type||Range (%)||Average Recovery (%)|
|EDTA plasma (n=5)||92-104||98|
|Cell culture media (n=5)||87-101||93|
Samples were spiked with high concentrations of Human ERK3 and diluted with Reference Standard & Sample Diluent to produce samples with values within the range of the assay.
|Serum (n=5)||EDTA plasma (n=5)||Cell culture media (n=5)|
An unopened kit can be stored at 4°C for 1 month. If the kit is not used within 1 month, store the items separately according to the following conditions once the kit is received.
|Micro ELISA Plate(Dismountable)||8 wells ×12 strips||-20°C, 6 months|
|Reference Standard||2 vials|
|Concentrated Biotinylated Detection Ab (100×)||1 vial, 120 µL|
|Concentrated HRP Conjugate (100×)||1 vial, 120 µL||-20°C(shading light), 6 months|
|Reference Standard & Sample Diluent||1 vial, 20 mL||4°C, 6 months|
|Biotinylated Detection Ab Diluent||1 vial, 14 mL|
|HRP Conjugate Diluent||1 vial, 14 mL|
|Concentrated Wash Buffer (25×)||1 vial, 30 mL|
|Substrate Reagent||1 vial, 10 mL||4°C(shading light)|
|Stop Solution||1 vial, 10 mL||4°C|
|Plate Sealer||5 pieces|
|Product Description||1 copy|
|Certificate of Analysis||1 copy|
- Set standard, test sample and control (zero) wells on the pre-coated plate and record theirpositions. It is recommended to measure each standard and sample in duplicate. Note: addall solutions to the bottom of the plate wells while avoiding contact with the well walls. Ensuresolutions do not foam when adding to the wells.
- Aliquot 100µl of standard solutions into the standard wells.
- Add 100µl of Sample / Standard dilution buffer into the control (zero) well.
- Add 100µl of properly diluted sample (serum, plasma, tissue homogenates and otherbiological fluids) into test sample wells.
- Cover the plate with the sealer provided in the kit and incubate for 90 min at 37°C.
- Aspirate the liquid from each well, do not wash. Immediately add 100µL of BiotinylatedDetection Ab working solution to each well. Cover the plate with a plate seal and gently mix. Incubate for 1 hour at 37°C.
- Aspirate or decant the solution from the plate and add 350µL of wash buffer to each welland incubate for 1-2 minutes at room temperature. Aspirate the solution from each well andclap the plate on absorbent filter paper to dry. Repeat this process 3 times. Note: a microplatewasher can be used in this step and other wash steps.
- Add 100µL of HRP Conjugate working solution to each well. Cover with a plate seal andincubate for 30 min at 37°C.
- Aspirate or decant the solution from each well. Repeat the wash process for five times asconducted in step 7.
- Add 90µL of Substrate Reagent to each well. Cover with a new plate seal and incubate forapproximately 15 min at 37°C. Protect the plate from light. Note: the reaction time can beshortened or extended according to the actual color change, but not by more than 30min.
- Add 50 µL of Stop Solution to each well. Note: Adding the stop solution should be done inthe same order as the substrate solution.
- Determine the optical density (OD value) of each well immediately with a microplate readerset at 450 nm.