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Mouse Dual specificity mitogen-activated protein kinase kinase 6 (Map2k6) ELISA Kit (MOEB1634)

SKU:
MOEB1634
Product Type:
ELISA Kit
Size:
96 Assays
Uniprot:
P70236
ELISA Type:
Sandwich
Reactivity:
Mouse
$779
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Description

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Mouse Dual specificity mitogen-activated protein kinase kinase 6 (Map2k6) ELISA Kit

The Mouse MAP2K6 (Dual Specificity Mitogen-Activated Protein Kinase Kinase 6) ELISA Kit is a reliable tool for detecting levels of MAP2K6 in mouse serum, plasma, and cell culture supernatants. This kit offers high sensitivity and specificity, ensuring accurate and consistent results for a variety of research applications.MAP2K6 is a key regulator of the MAPK signaling pathway, playing a crucial role in cell growth, differentiation, and response to external stimuli.

Dysregulation of MAP2K6 has been implicated in various diseases, including cancer, inflammatory disorders, and neurodegenerative diseases. Thus, this ELISA kit is essential for studying the function and potential therapeutic targeting of MAP2K6 in these pathological conditions.Overall, the Mouse MAP2K6 ELISA Kit is a valuable tool for researchers looking to investigate the role of MAP2K6 in disease pathogenesis and explore novel therapeutic strategies.

Product Name:Mouse Dual specificity mitogen-activated protein kinase kinase 6 (Map2k6) ELISA Kit
SKU:MOEB1634
Size:96T
Target:Mouse Dual specificity mitogen-activated protein kinase kinase 6 (Map2k6)
Synonyms:MAPK/ERK kinase 6, SAPKK3, MEK 6, MAP kinase kinase 6, Prkmk6, Sapkk3
Assay Type:Sandwich
Detection Method:ELISA
Reactivity:Mouse
Detection Range:0.156-10ng/mL
Sensitivity:0.093ng/mL
Intra CV:Provided with the Kit
Inter CV:Provided with the Kit
Linearity:Provided with the Kit
Recovery:Provided with the Kit
Function:Dual specificity protein kinase which acts as an essential component of the MAP kinase signal transduction pathway. With MAP3K3/MKK3, catalyzes the concomitant phosphorylation of a threonine and a tyrosine residue in the MAP kinases p38 MAPK11, MAPK12, MAPK13 and MAPK14 and plays an important role in the regulation of cellular responses to cytokines and all kinds of stresses. Especially, MAP2K3/MKK3 and MAP2K6/MKK6 are both essential for the activation of MAPK11 and MAPK13 induced by environmental stress, whereas MAP2K6/MKK6 is the major MAPK11 activator in response to TNF. MAP2K6/MKK6 also phosphorylates and activates PAK6. The p38 MAP kinase signal transduction pathway leads to direct activation of transcription factors. Nuclear targets of p38 MAP kinase include the transcription factors ATF2 and ELK1. Within the p38 MAPK signal transduction pathway, MAP3K6/MKK6 mediates phosphorylation of STAT4 through MAPK14 activation, and is therefore required for STAT4 activation and STAT4-regulated gene expression in response to IL-12 stimulation. The pathway is also crucial for IL-6-induced SOCS3 expression and down-regulation of IL-6-mediated gene induction; and for IFNG-dependent gene transcription. Has a role in osteoclast differentiation through NF-kappa-B transactivation by TNFSF11, and in endochondral ossification and since SOX9 is another likely downstream target of the p38 MAPK pathway. MAP2K6/MKK6 mediates apoptotic cell death in thymocytes. Acts also as a regulator for melanocytes dendricity, through the modulation of Rho family GTPases.
Uniprot:P70236
Sample Type:Serum, plasma, tissue homogenates, cell culture supernates and other biological fluids
Specificity:Natural and recombinant mouse Dual specificity mitogen-activated protein kinase kinase 6
Sub Unit:Dimer. Interacts (via its D domain) with its substrates MAPK11, MAPK12, MAPK13 and MAPK14. Interacts (via its DVD domain) with MAP3Ks activators like MAP3K5/ASK1, MAP3K1/MEKK1, MAP3K2/MEKK2, MAP3K3/MEKK3, MAP3K4/MEKK4, MAP3K7/TAK1, MAP3K11/MLK3 and MAP3K17/TAOK2. Interacts with DCTN1. Interacts with EIF2AK2/PKR.
Research Area:Cancer
Subcellular Location:Nucleus Cytoplasm Cytoplasm Cytoskeleton Binds to microtubules.
Storage:Please see kit components below for exact storage details
Note:For research use only
UniProt Protein Function:MKK6: a dual-specificity protein kinase of the STE7 family. Activates p38 MAP kinase by phosphorylating a Thr and a Tyr residue in the activation loop. Is activated by cytokines and environmental stress in vivo. Three alternatively spliced isoforms have been reported.Protein type: Protein kinase, dual-specificity (non-receptor); Protein kinase, STE; EC 2.7.12.2; Kinase, protein; STE group; STE7 familyCellular Component: cytoplasmMolecular Function: MAP kinase kinase activity; protein binding; protein kinase binding; receptor signaling protein serine/threonine kinase activityBiological Process: activation of MAPK activity; cardiac muscle contraction; MAPKKK cascade; positive regulation of apoptosis; positive regulation of nitric-oxide synthase biosynthetic process; positive regulation of prostaglandin secretion; positive regulation of protein amino acid phosphorylation; protein amino acid phosphorylation
UniProt Protein Details:
NCBI Summary:
UniProt Code:P70236
NCBI GenInfo Identifier:3024165
NCBI Gene ID:26399
NCBI Accession:P70236.1
UniProt Secondary Accession:P70236
UniProt Related Accession:P70236
Molecular Weight:37,432 Da
NCBI Full Name:Dual specificity mitogen-activated protein kinase kinase 6
NCBI Synonym Full Names:mitogen-activated protein kinase kinase 6
NCBI Official Symbol:Map2k6
NCBI Official Synonym Symbols:MEK6; MKK6; Prkmk6; SAPKK3
NCBI Protein Information:dual specificity mitogen-activated protein kinase kinase 6
UniProt Protein Name:Dual specificity mitogen-activated protein kinase kinase 6
UniProt Synonym Protein Names:MAPK/ERK kinase 6; MEK 6; SAPKK3
Protein Family:Dual specificity mitogen-activated protein kinase kinase
UniProt Gene Name:Map2k6
UniProt Entry Name:MP2K6_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.

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