Bovine il 1β elisa reagent kit

Production of IL-1β, IL-6, TNF-α, and CXCL8 (IL-8) from both the apical and basal surfaces of infected and uninfected (PBS alone added to the apical surface) BBEC cultures was assessed at each time point p.i. To measure cytokine/chemokine production from the basal surface, 1 ml of medium was removed from the basolateral compartment and centrifuged at 5,000 × g for 5 min, and the supernatant was immediately frozen at −80°C. To measure levels of cytokine/chemokine production from the apical surface, 0.5 ml of antibiotic-free ALI medium was added to the apical surface of each culture and the cultures were returned to the incubator for 30 min. The medium was subsequently removed and centrifuged at 5,000 × g for 5 min, and the supernatant was immediately frozen at −80°C. Cytokine/chemokine production was quantified using commercially available enzyme-linked immunosorbent assay (ELISA) kits according to the instructions of the manufacturers as follows: IL-1β, bovine IL-1β ELISA reagent kit (Thermo Fisher; catalog no. ESS0027); IL-6, bovine IL-6 ELISA reagent kit (Thermo Fisher; catalog no. ESS0029); CXCL8, bovine IL-8 ELISA development kit (Mabtech; catalog no. 3114-1A-6); TNF-α, bovine TNF-α DuoSet ELISA development system (R&D systems; catalog no. DY2279). Triplicate samples were measured for each insertion, and two individual cultures were analyzed for each donor animal (n = 6).