Minimum essential medium eagle

The presence of anti-vimentin and anti-actin antibodies was determined using indirect immunofluorescence on monolayers of colchicine-treated Hep2 cells, as described elsewhere [11 (link), 12 (link)]. Briefly, Hep2 monolayer cells culture was home-prepared. This culture performed on slide was incubated with colchicine 0.0014 % (w:v) (Sigma) diluted in minimum essential medium Eagle (Eurobio), glutamine supplemented, during 20 h at 37 °C. After three washes with phosphate-buffered saline (PBS), and acetone-fixed, the monolayer cells was incubated with sera at the dilution of 1/40, in PBS, during 30 min. After PBS washing (3×), monolayer cells were revealed using a fluorochrome–labeled, polyclonal antihuman IgG, IgA, IgM antiserum (BioRad Laboratory), 30 min incubated. Vimentin appear to be collapsed into thick perinuclear coils if the tested serum was positive for anti-vimentin antibodies, and a typical pattern of actin cables was strongly stained if serum was positive for anti-actin antibodies.

Human recombinant TGF-β1 was obtained from PeproTech, (Cranbury, NJ, USA). FICZ and kynurenine were purchased from Sigma (St. Louis, MO, USA). ITE and curcumin were purchased from TOCRIS (Bristol, UK). Fetal bovine serum (FBS) was purchased from Gibco (Waltham, MA, USA) and penicillin, streptomycin, L-glutamine from Dutscher (Brumath, France). MEM non-essential amino acid and Cellytic™ buffer were supplied by Sigma (St. Louis, MO, USA). Minimum Essential Medium Eagle was obtained from Eurobio (Montpellier, France).