Corticosterone plasma levels
The plasma was obtained in first day and in 7th day
post-stress induction after whole blood centrifugation at 3000 xg at
room temperature for 10 minutes and stored at -80 ºC until analysis. The
corticosterone circulating level was measured employing an ELISA Kit
(Enzo Life Sciences, Farmingdale, New York, USA) following the product
manual. Briefly, the samples were diluted according to protocol. Then,
100 µl of the sample or standard solutions were incubated at the plate
with the antibody provided by the kit at room temperature on a plate
shaker for 2 hours. After, the wells were empty and washed and added 200
µl of pNpp Substrate Solution incubating for 1 hour without shaking.
Finally, the plate was read at 405 nm using a microplate reader
(SpectraMax I3; Molecular Devices,
San Jose, California, USA).