Endothelial NOS (eNOS), also known as nitric oxide synthase 3 (NOS3) or constitutive NOS (cNOS), is an enzyme that in humans is encoded by the NOS3 gene located in the 7q35-7q36 region of chromosome 7. This enzyme is one of three isoforms that synthesize nitric oxide (NO), a small gaseous and lipophilic molecule that participates in several biological processes. The other isoforms include neuronal nitric oxide synthase (nNOS), which is constitutively expressed in specific neurons of the brain and inducible nitric oxide synthase (iNOS), whose expression is typically induced in inflammatory diseases. eNOS is primarily responsible for the generation of NO in the vascular endothelium, a monolayer of flat cells lining the interior surface of blood vessels, at the interface between circulating blood in the lumen and the remainder of the vessel wall. NO produced by eNOS in the vascular endothelium plays crucial roles in regulating vascular tone, cellular proliferation, leukocyte adhesion, and platelet aggregation. Therefore, a functional eNOS is essential for a healthy cardiovascular system.
Background References
1. Yao L et al. Non-steroidal mineralocorticoid receptor antagonist finerenone ameliorates mitochondrial dysfunction via PI3K/Akt/eNOS signaling pathway in diabetic tubulopathy. Redox Biol. 2023 Dec
2. Teng Z et al. eNOS polymorphisms on male infertility: An updated systematic review and meta-analysis. Medicine (Baltimore). 2023 Jun
Western blot analysis of eNOS on different lysates with Rabbit anti-eNOS antibody (HA724143) at 1/2,000 dilution.
Lane 1: HeLa cell lysate (negative) (20 µg/Lane) Lane 2: EA.hy926 cell lysate (20 µg/Lane) Lane 3: RAW264.7 cell lysate (negative) (20 µg/Lane) Lane 4: Mouse kidney tissue lysate (no heat) (20 µg/Lane)
Notice: no heat means the lysate is not boiled.
Predicted band size: 133 kDa Observed band size: 133 kDa
Exposure time: 6 seconds; ECL: K1801;
4-20% SDS-PAGE gel.
Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA724143) at 1/2,000 dilution was used in primary antibody dilution (K1803) at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.
eNOS was immunoprecipitated from 0.2 mg EA.hy926 cell lysate with HA724143 at 2 µg/10 µl beads. Western blot was performed from the immunoprecipitate using HA724143 at 1/1,000 dilution. HRP Conjugated Anti-Rabbit IgG for IP Nano-secondary antibody at 1/5,000 dilution was used for 1 hour at room temperature.
Lane 1: EA.hy926 cell lysate (input) Lane 2: HA724143 IP in EA.hy926 cell lysate Lane 3: Rabbit IgG instead of HA724143 in EA.hy926 cell lysate