Description: This CXNFT monoclonal antibody reacts to mouse NOS2 (inducible NOS, iNOS). Nitric oxide synthase enzymes catalyze the formation of nitric oxide from L-arginine through an NADPH- and oxygen-dependent mechanism. There are three isoforms of NOS that are encoded by three separate genes. NOS1 (neuronal NOS, nNOS) and NOS3 (endothelial NOS, eNOS) are constitutively expressed, while NOS2 is induced in response to bacterial endotoxins and inflammatory cytokines such as IFN gamma and TNF alpha. NOS2 is expressed by myeloid-derived suppressor cells and M1 macrophages but not alternatively activated M2 macrophages. NOS enzymes are functionally active only when they form homodimers, and dimerization of NOS2 occurs at steady-state concentrations of free Ca2+ such that NOS2 is functionally active when it is produced. Applications Reported: This CXNFT antibody has been reported for use in intracellular staining followed by flow cytometric analysis. Applications Tested: This CXNFT antibody has been tested by intracellular staining and flow cytometric analysis of stimulated mouse thioglycolate-elicited peritoneal exudate cells using the intracellular Fixation and Permeabilization Buffer Set (cat. 88-8824) and protocol. The Foxp3/Transcription Factor Staining Buffer Set (cat. 00-5523) may also be used with similar results. This can be used at less than or equal to 0.06 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. It is recommended that the antibody be carefully titrated for optimal performance in the assay of interest. PerCP-eFluor™ 710 emits at 710 nm and is excited with the blue laser (488 nm); it can be used in place of PerCP-Cyanine5.5. We recommend using a 710/50 bandpass filter, however, the 695/40 bandpass filter is an acceptable alternative. Please make sure that your instrument is capable of detecting this fluorochrome. 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) (100 μL of cell sample + 100 μL of IC Fixation Buffer) or 1-step Fix/Lyse Solution (Product No. 00-5333) 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 specific performance should be determined empirically. Excitation: 488 nm; Emission: 710 nm; Laser: Blue Laser. Filtration: 0.2 μm post-manufacturing filtered. iNOS (Inducible Nitric oxide, NO, NOS) is an inorganic, gaseous free radical that carries a variety of messages between cells. Vasorelaxation, neurotransmission and cytotoxicity can all be potentiated through cellular response to NO. NO production is mediated by members of the nitric oxide synthase (NOS) family. iNOS is expressed in liver and inducible by a combination of lipopolysaccharide and certain cytokines. NOS catalyzes the oxidization of L-arginine to produce L-citrulline and NO. Two constitutive isoforms, brain or neuronal NOS (b or nNOS, type I) and endothelial cell NOS (eNOS, type III), and one inducible isoform (iNOS, type II), have been cloned. All NOS isoforms contain calmodulin, nicotinamide adenine dinucleotide phosphate (NADPH), flavin adenine dinucleotide (FAD), and flavin mononucleotide (FMN) binding domains. iNOS is found in a variety of cell types including macrophages, hepatocytes, synoviocytes, and smooth muscle cells. Cytokines such as interferon-gamma (IFN), tumor necrosis factor (TNF), interleukin-1 and -2, and lipopolysaccarides (LPS) cause an increase in iNOS mRNA, protein, and activity levels. Protein kinase C-stimulating agents exhibit the same effect on iNOS activity. After cytokine induction, iNOS exhibits a delayed activity response which is then followed by a significant increase in NO production over a long period of time. Three related iNOS pseudogenes are located within the Smith-Magenis syndrome region on chromosome 17. Diseases associated with iNOS dysfunction include achalasia and impotence.
|4° C, store in dark, DO NOT FREEZE!|
|PBS with 0.1% gelatin and 0.09% sodium azide; pH 7.2|