Antibody arrays

We measured the relative concentrations of a total of 170 proteins (see Additional file 1, Supplementary Table S2) with antibody arrays (RayBiotech Inc., Peachtree Corners, GA, USA) according to the manufacturer’s instructions. Briefly, a custom glass-based antibody array targeting the 170 proteins of interest was built, 100 μl of diluted serum sample was added to each well, incubated overnight at 4°C, and then extensively washed. We then incubated the wells with biotin-conjugated antibodies specific to the different proteins. Membranes were developed with Alexa Fluor^® 555-conjugated streptavidin (Thermo Fisher Scientific, Waltham, MA, USA). The slides were scanned with 532 nm excitation and extracted using an InnoScan^® 300 Scanner (Innopsys, Carbonne, France).

IRISOE1-5 grown in the absence of Dox (i.e. naïve HME) and in the presence of Dox (i.e. IRISOE1-5 or HME/IRIS) cells were assessed for differential factors. Briefly, conditioned media (CM) from equal number of either cell line plated in serum-free medium for 20h under standard conditions was used to screen cytokine, chemokine and growth factors antibody arrays (RayBio, Norcross, GA, USA) performed according to the manufacturer’s instructions and as described previously [74 (link)].