G2000

CM (1% FBS) from NCF or ILβ-treated NCF or IL1β-treated NCF plus neutralizing IL1β antibody was used for hybridization on the glass-slide human cytokine array G2000 from RayBiotech (Norcross, GA, USA), which detects 174 human cytokines. Samples were processed and analyzed by Tebu-Bio (Le Perray-en-Yvelines, France).
IL6 and IL1β were determined in supernatants obtained from cocultures CM by means of ELISA (R&D cat number D6050 and DLB50, respectively).

CM was freshly collected as described above from chemotherapy-treated human and mouse fibroblasts or from control fibroblasts. CM was collected independently from two flasks per condition, and 100 μL added to duplicate RayBiotech G2000 array slides. Wash and incubation steps were performed according to the manufacturer’s protocol. Fluorescence signal detection at 532 nm was performed on a GenePix 4000B array scanner. Median fluorescence intensity (MFI) was quantified using ImageJ software. Fold changes were calculated by dividing the mean MFI for the two chemotherapy-treated CM replicates by the mean MFI for the two control replicates for each fibroblast type.