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%.