Figure 1 In vitro validation of promoter and transgene function. Fibroblast‐like synoviocytes (FLS) were transduced with rAAV5 containing the NFκB.TNFRI‐Ig or the CMV‐TNFRI‐Ig gene. After 24 hours, interleukin (IL)1β (100 ng/ml), tumour necrosis factor (TNF)α (100 ng/ml) or lipopolysaccharide (LPS; 1 μg/ml) was added to the culture medium in the presence or absence of the nuclear factor kappa B (NFκB) blocking agent pyrrolidinedithiocarbamate (PDTC; 200 μmol/l). (A) Forty‐eight hours later, the supernatants were collected and an ELISA was performed to analyse the amount of TNFRI‐Ig protein. The capacity of produced TNFRI‐Ig to bind and neutralise human and rat TNFα was tested in a TNFα bioassay. (B) The percentage of TNFα neutralisation by TNFRI‐Ig is directly related to the percentage of the number of surviving cells and expressed as percentage of TNFα blocking. The presented results are performed in triplicate and are representative for two independent experiments.