Phospho-p38 MAPK (Thr180, Tyr182) Monoclonal Antibody (4NIT4KK), PE-Cyanine7, eBioscience™, Invitrogen™

Beschreibung

Description: This 4NIT4KK monoclonal antibody recognizes human and mouse p38 mitogen-activated protein kinase (MAPK) when phosphorylated on T180/Y182. p38 MAPK belongs to a family of conserved serine/threonine protein kinases that are phosphorylated and activated in response to numerous stress stimuli including TLR ligands (such as LPS), osmotic shock, heat shock, UV irradiation, and inflammatory cytokines. There are four p38 MAPK members that include p38 alpha, p38 beta, p38 gamma, and p38 delta. The primary activators of p38 MAPK are MKK3/4 and MKK6. Several downstream targets of p38 MAPK have been identified including MK2/3, p53, ATF-2, and ETS1. p38 MAPK can be negatively regulated by the chemical inhibitor SB203580. Specificity of this 4NIT4KK clone was determined by ELISA, flow cytometry, and western blotting. Applications Reported: This 4NIT4KK antibody has been reported for use in intracellular staining followed by flow cytometric analysis. Applications Tested: This 4NIT4KK antibody has been pre-diluted and tested by intracellular staining followed by flow cytometric analysis of stimulated normal human peripheral blood cells. This may be used at 5 μL (0.25 μg) per test. A test is defined as the amount (μg) of antibody that will stain a cell sample in a final volume of 100 μL. Cell number should be determined empirically but can range from 10^5 to 10^8 cells/test. Staining Protocol: All protocols work well for this monoclonal antibody.

Use of Protocol A: Two-step protocol: intracellular (cytoplasmic) proteins allows for the greatest flexibility for detection of surface and intracellular (cytoplasmic) proteins. Use of Protocol B: One-step protocol: intracellular (nuclear) proteins is recommended for staining of transcription factors in conjunction with surface and phosphorylated intracellular (cytoplasmic) proteins. Protocol C: Two-step protocol: Fixation/Methanol allows for the greatest discrimination of phospho-specific signaling between unstimulated and stimulated samples, but with limitations on the ability to stain specific surface proteins (refer to "Clone Performance Following Fixation/Permeabilization" located in the Best Protocols Section under the Resources tab online). All Protocols can be found in the Flow Cytometry Protocols: "Staining Intracellular Antigens for Flow Cytometry Protocol" located in the Best Protocols Section under the Resources tab online. Light sensitivity: This tandem dye is sensitive to photo-induced oxidation. Please protect this vial and stained samples from light. Fixation: Samples can be stored in IC Fixation Buffer (Product No. 00-8222-49) (100 μL of cell sample + 100 μL of IC Fixation Buffer) or 1-step Fix/Lyse Solution (Product No. 00-5333-57) for up to 3 days in the dark at 4°C with minimal impact on brightness and FRET efficiency/compensation. Some generalizations regarding fluorophore performance after fixation can be made, but clone specificperformance should be determined empirically. Excitation: 488-561 nm; Emission: 775 nm; Laser: Blue Laser, Green Laser, Yellow-Green Laser Mitogen-activated protein kinase 11.The protein encoded by this gene is a member of the MAP kinase family. MAP kinases act as an integration point for multiple biochemical signals, and are involved in a wide variety of cellular processes such as proliferation, differentiation, transcription regulation, and development. This kinase is most closely related to p38 MAP kinase, both of which can be activated by proinflammatory cytokines and environmental stress. This kinase is activated through its phosphorylation by MAP kinase kinases (MKKs), preferably by MKK6. Transcription factor ATF2/CREB2 has been shown to be a substrate of this kinase.