CGRP assay
Firstly, affinity sorbent (A19482, SPI Bio, Bertin Pharma) was used with a pool of different sources of plasma to prepare CGRP-free plasma. Then, the samples were analyzed using a commercially available kit by following the manufacturer’s procedures: CGRP (A05482, SPI Bio, Bertin Pharma). The concentration of CGRP is measured using Ellman’s Reagent to detect the enzymatic activity of the AChE. The intensity of the yellow color formed is proportional to the amount of CGRP present in that sample [11]. The absorbance was read at 405 nm on SpectraMax I3 (Molecular Devices, San Jose, California, USA). The absorbance values of the standards were used to plot a standard curve, from which absorbance values of experimental samples were interpolated to determine their concentrations. The sensitivities of the assay were 2 pg/ml using quality control. This test was performed in mice plasma of 7th-day post-stress induction.