Description
Nitric oxide synthase (NOS), a cell-type specific enzyme, catalyzes the synthesis of nitric oxide (NO). NO is a short-lived radical that transmits signals involved in vasorelaxation, neurotransmission, and cytotoxicity. In neurons and endothelial cells, constitutive NOS (cNOS) is activated by agonists that increase intracellular Ca2+ levels and enhance calmodulin binding. Neuronal NOS (nNOS) and endothelial NOS (eNOS) have recognition sites for NADPH, FAD, FMN, and calmodulin and both are regulated in a similar manner. The human forms exhibit 52% amino acid identity. However, they are distinct gene products of about 155 kDa (nNOS) and 140 kDa (eNOS). The eNOS gene was cloned from human vascular endothelium as well as from bovine aortic endothelial cells (BAEC). eNOS protein has a unique N-myristylation consensus sequence that may explain its membrane localization. The 33/eNOS monoclonal antibody recognizes eNOS, regardless of phosphorylation status.
Either BD Cytofix™ fixation buffer or BD Phosflow™ Fix Buffer I may be used for cell fixation.
Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze. The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography. The antibody was conjugated to Alexa Fluor® 647 under optimum conditions, and unreacted Alexa Fluor® 647 was removed.
1. This reagent has been pre-diluted for use at the recommended Volume per Test. We typically use 1 × 10^6 cells in a 100-µl experimental sample (a test).
2. Alexa Fluor® 647 fluorochrome emission is collected at the same instrument settings as for allophycocyanin (APC).
3. The Alexa Fluor®, Pacific Blue™, and Cascade Blue® dye antibody conjugates in this product are sold under license from Molecular Probes, Inc. for research use only, excluding use in combination with microarrays, or as analyte specific reagents. The Alexa Fluor® dyes (except for Alexa Fluor® 430), Pacific Blue™ dye, and Cascade Blue® dye are covered by pending and issued patents.
4. Caution: Sodium azide yields highly toxic hydrazoic acid under acidic conditions. Dilute azide compounds in running water before discarding to avoid accumulation of potentially explosive deposits in plumbing.
5. Source of all serum proteins is from USDA inspected abattoirs located in the United States.
6. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
7. Alexa Fluor® is a registered trademark of Molecular Probes, Inc., Eugene, OR.
8. An isotype control should be used at the same concentration as the antibody of interest.
9. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
