Leukotriene b4 eia kit

Samples were analysed for leukotriene B4 and 8-isoprostane concentrations using enzyme-linked immune-assays (ELISA). Any samples with insufficient volume for both assays were prioritised for 8-isoprostane analysis. ELISA buffer, ELISA standards, samples, AChE tracers and ELISA antiserums for leukotriene B4 and 8-isoprostane were added to the assay plates according to the protocol outlined by the Cayman Chemical Leukotriene B4 EIA Kit (Item No. 520,111) and 8-Isoprostane EIA Kit (Item No. 516,351). A standard curve was plotted using the absorbance readings of the maximum binding, non-specific binding and standard wells. The standard curve was then used to determine the concentrations of leukotriene B4 and 8-isoprostane in the EBC samples. Measures below the limit of detection were assigned a concentration of 0 pg/mL.

Plasma cytokine levels (IL-2, IL-4, IL-5, IL-6, IL-10, IL-12, IL-13, 1L-17A, IL-23, IFN-γ, TNF-α, and TGF-β1) were determined using a Multi-Analyte Profiler ELISArray® (Superarray Bioscience Corporation, CAT#1026 A). Eicosanoid levels were assessed employing the acetylcholinesterase-based assay PGE₂ EIA kit and leukotriene B₄ EIA kit (Cayman Chemical, CAT#514531).

Transiently transfected A549 cells were incubated in serum-free media and stimulated with 4 μM calcium ionophore (A23817, Sigma) for 30 min prior to collecting supernatants [61 (link)]. H1299 Tet-On cells were incubated in serum-free media containing 10 μM arachidonic acid (Cayman Chemical) for 30 min. Supernatants were removed from the cells and centrifuged at 2,000 x g, 10 min, 4°C. These cell-free supernatants were then analyzed using the Leukotriene B4 EIA Kit (Cayman Chemical) according to the manufacturer's provided protocol. The data represent the average of three independent experiments. Each sample was measured with n ≥ 2 technical replicates.