2.8 Multiplex analysis of cytokines
Quantification of IL-6 and IL-1β cytokines in hippocampal samples was performed as described previously (Sanchez-Alavez, Nguyen et al., 2019) by a multiplexed flow cytometric assay using a rat cytokine kit (Millipore, Billerica, MA, USA, Cat# RECYTMAG-65K) according to the manufacturer’s instructions. Briefly, hippocampal tissue lysate and pre-mixed antibody beads were incubated at room temperature for 2 h, washed three times, and incubated with streptavidin-phycoerythrin for 30 min. The contents were washed and resuspended in sheath fluid before being read on the Luminex MAGPIX system. The results were analysed using MILLIPLEX™ Analyst 5.1 software (Luminex Corporation, Austin, TX, USA). The concentrations of cytokines were determined based on standard curves, and the coefficient of variation was calculated; it did not exceed 20%.