sFIS Assay for Interferon-Stimulated Genes

This sFIS assay [36 (link)] consisted of THP1-Dual™ myeloid reporter cells (InvivoGen, Toulouse, France) seeded in a 96-well plate at a density of 30,000 cells per well in 100 µL media (RPMI-1640 containing 2 mM L-glutamine, 25 mM HEPES, 100 units/mL penicillin, 100 µg/L streptomycin, 100 µg/mL Normocin and 10% heat inactivated fetal calf serum). After 24 h, THP1 cells were treated with 100 µL of patient plasma for 24 h. To determine activation of interferon-stimulated response elements (ISRE) coding for interferon (IFN)-stimulated genes (ISG), luciferase was checked in media by adding 50 µL of Quanti-Luc GOLD (InvivoGen) to 100 µL of (separately recovered) THP1 media. Bioluminescence was examined for 100 ms immediately after Quanti-Luc GOLD addition by microplate reader (Agilent, Santa Clara, CA, USA). IFN/ISG response activity values were normalized using the results of samples taken before RT treatment of each individual patient.