Mouse il 6 duoset kit

Culture supernatants were collected from mouse primary microglia exposed to various treatments and assessed for expression of IL‐6 / TNFα protein using the mouse IL‐6 DuoSet Kit or mouse TNF‐α DuoSet Kit (R&D Systems, Minneapolis, MN).

To measure secreted IL-6, conditioned medium was collected from each sample collected above. IL-6 was quantified with the mouse IL-6 DuoSet kit from R&D Systems according to the manufacturer’s instructions. Plates were washed with PBS at pH 7.4 containing 0.05% Tween-20 (PBS-T) and blocked using PBS containing 1% IgG-free BSA (Jackson Immunological West Grove, PA). The substrate used was Thermo Pierce (Rockford, IL) 1-Step Ultra TMB. A seven point standard curve was prepared by serial dilution of the included 1000 pg/mL standard. Sample concentrations were interpolated from a 4 parameter logistic fit of the standards. All samples and standards were assayed in duplicate. All ELISA analyses had a sample size of three.