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Protocol |
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Control peptide should be coated at 1 ug/ml. Control peptide, because of its low mol. Wt (<3 kDa), is not suitable for Western. It should be used for ELISA or antibody blocking to confirm antibody specificity
Human, rat, mouse, and bovine eNOS/NOS-3 are ~1202-1205 aa proteins (1). A 15 amino acid peptide sequence corresponding to human eNOS (designated eNOS-P; control peptide; 596-610 aa) (1) was synthesized and coupled to KLH. Control peptide was also used for affinity purification of antibodies.
Human eNOS-P peptide sequence is 100%conserved in mouse/bovine (15/15 aa), 93% in rat (14/15 aa) and 86% in pig (13/15 aa). No significant sequence homology of eNOS-P is seen with NOS-1/NOS-2 or other proteins.
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Substrate/Buffer |
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PBS, pH 7.4 at 1 mg/ml (100 ug/100 ul)
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Storage |
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peptide received in solution or reconstituted from lyophilized vials should be stored frozen at -20oC or below in suitable aliquots. It is not recommended to store diluted solutions. Avoid repeated freeze and thaw
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References |
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1.MARSDEN P.A FEBS LETT. 307, 287-293 (1992); JANSSENS S.P., J. BIOL. CHEM. 267, 14519-14522 (1992); Michel T and Lamas, S (1992) J Card. Pharmacol.20, S45-S49
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Notes |
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NO is synthesized by L-arginine, oxygen, and NADPH by three known isoforms of heme-containing flavoproteins termed NO synthase (NOS, I-III, mol wt. ~130-160 kDa). One group of enzyme is constitutive, agonist-triggered, and dependent on Ca2+/Calmodulin and is inhibited by L-arginine analogues (L-NNA, L-NMMA). It is found in endothelium, adrenal glands, brain and platelets. The other principle group is inducible, Ca2+/Calmodulin-independent, and inhibited by NMMA and L-NNA. It has been found in macrophage, hepatocytes, tumor cells, vascular smooth muscle and endothelial cells. Analyses of cDNA clones have identified three distinct NOS genes in mammals: neuronal (nNOS/bNOS/NOS-I), endothelial (eNOS/NOS-III), and macrophage (mNOS/iNOS/NOS-II). Both nNOS and eNOS are constitutive and the mNOS/iNOS is inducible. Sequence homology among different cloned isoforms is ~ 50%.
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