Omega mars analysis software

H₂O₂ generation was measured as previously described (32 (link)). In brief, neutrophils were cultured under normoxia or hypoxia for 1 h before addition of HRP (Sigma, UK) and Amplex^® UltraRed (Invitrogen, UK). H₂O₂ generation in response to phorbol 12-myristate 13-acetate (PMA) (Sigma, UK) was recorded using a FLUOstar Omega microplate reader (BMG Labetech, Germany) and rates (expressed in nM/sec) determined using Omega Mars Analysis software (BMG Labtech, Germany).

2 × 10⁶ HC-neutrophils were incubated with 200 μg/ml IgG for 1 hour at 37 °C before addition of 0.5U/ml HRP (Sigma, UK) and 60 nM Amplex UltraRed (Invitrogen, UK). 4 × 10⁵ cells were added to each test well of a 96-well black microplate (Thermo Scientific, UK) and fluorescence measured by FLUOstar Omega microplate reader (BMG Labetech, Germany). H₂O₂ generation in response to 50 nM PMA was recorded over time and rates (expressed in nM/sec) determined using Omega Mars Analysis software (BMG Labtech, Germany). In separate experiments, H₂O₂ generation was assessed in patient- and HC-derived neutrophils, however in the absence of autologous IgG.