Western blot analysis of Atrial Natriuretic Peptide was performed by loading 25 µg of rat heart cell lysate onto an SDS polyacrylamide gel. Proteins were transferred to a PVDF membrane and blocked at 4ºC overnight. The membrane was probed with a Atrial Natriuretic Peptide polyclonal antibody (Product # PA3-228) at a dilution of 1:1000 overnight at 4°C, washed in TBST, and probed with an HRP-conjugated secondary antibody for 1 hr at room temperature in the dark. Chemiluminescent detection was performed using Pierce ECL Plus Western Blotting Substrate (Product # 32132). Results show a band at ~17 kDa.