Differential cell counting
To differentiate the various cell populations in the BALF,
cytocentrifuged cell preparations (Shandon Cytospin 3, 400 RPM, 10 min)
were used, with ~5x104 cells per
slide. Dried cytospots were subjected to a Shandon
Kwik-Diff® Fixative (Thermo Fischer Scientific, USA),
followed by Hemacolour® eosin and thiazine
differential stains (Merck, USA) as outlined in the manufacturer’s
instructions. Cell types (i.e. macrophages, neutrophils and lymphocytes)
were identified according to standard morphological criteria, using the
above-mentioned microscope, with at least 500 cells counted per slide.