... agents were added 60 min before the challenge with lipopolysaccharides (LPS) (10 ng/ml, Escherichia coli 055:B5; List Biological Laboratories ...
Efferocytosis was assessed by flow cytometry . Briefly, 5 × 105/well monocyte-derived human M2(IL-4+IL-10+Dex) macrophages or M2(Dex) macrophages were preincubated with 10 nM recombinant PA (List Biological Laboratories, Campbell, CA, USA), 10 nM recombinant EF (List Biological Laboratories), or various concentrations of PA + EF (ET) for 4 h at 37 °C in 5% CO2 atmosphere. Fluorescently-labeled apoptotic human PMN were preincubated with 10% serum, Protein S, or MFGE8 for 1 h unless otherwise noted and then added to the macrophages, resulting in a macrophage:PMN ratio of 1:5. After 1 h at 37 °C in 5% CO2 atmosphere, the percentage macrophages containing intracellular (eFluor 670+) but not surface-bound (surface CD66b−) PMN was assessed by flow cytometry.
Author did not specify which List Labs Anthrax Edema Factor was utilized. List Labs provides Product #178A (Anthrax Edema Factor (EF), Recombinant from B. anthracis) and Product #173B (Anthrax Edema Factor (EF), Recombinant, Mutant S447N from B. anthracis).