Biotinylated detection mabs

For the detection of cytokines in the cell culture supernatants or sera, ELISAs were performed as described previously [37 (link)], using the primary capture mAbs anti-TNFα, and anti-IL-6 and their corresponding biotinylated detection mAbs (Biolegend, USA). Recombinant mouse TNFα and IL-6 (Biolegend, USA) were used as standards. IL-10 was measured using a commercial ELISA kit (R&D systems, USA) according to manufacturer’s instructions. Other ELISA reagents included: HRP-conjugated avidin D (Vector Laboratories, USA), TMB microwell peroxidase substrate and stop solution (Kirkegaard and Perry Laboratories, USA).

For the detection of cytokines in the cell culture supernatants or sera, ELISAs were performed as described previously(17 (link)), using the primary capture mAbs anti-TNFα, anti-IL-6, and anti-IL-1β and their corresponding biotinylated detection mAbs (Biolegend, San Diego, CA). Other ELISA reagents included: recombinant mouse TNFα, IL-6, and IL-1β as standards (Biolegend, San Diego, CA), HRP-conjugated avidin D (Vector Laboratories, Burlingame, CA), and TMB microwell peroxidase substrate and stop solution (Kirkegaard and Perry Laboratories, Gaithersburg, MD). IL-10 cytokine was measured using Mouse IL-10 DuoSet ELISA kit (R&D Systems, Minneapolis, MN) according to manufacturer’s instructions. Biological replicates were used, representing independent BMDM cultures from individual mice.