Centroliapc lb 692 plate luminometer

The reactive oxygen species (ROS) assay was carried out as described (Huang et al., 2013 (link)). Briefly, nine 0.25 cm² leaf disks were excised from fully expanded leaves from 5-week-old Arabidopsis plants. The disks were incubated overnight in a 96-well plate with 100 µl of sterile water. Water was then replaced by 100 µl reaction solution [2 µl luminol (Sigma), 10 µg/ml horseradish peroxidase (Sigma)], with 100 nM flg22 or water (mock). The plate was analyzed at the indicated intervals for a period of 30 min using a CentroLIApc LB 692 plate luminometer [Berthold Technologies, (Bad Wildbad, Germany)].

Reactive Oxygen Species (ROS) evaluation was performed as previously described (Huang et al., 2012 (link)). Eight leaf discs of 0.25 cm² from three 5-week-old Arabidopsis plants were incubated in 100 μL ddH₂O overnight in 96-well plates. Water was then replaced by 100 μL of reaction solution containing 50 μM of luminol and 10 μg/mL of horseradish peroxidase (Sigma) supplemented with 50 nM of flg22 or water only for the mock controls. ROS measurements expressed as means of RLU (Relative Light Units) were conducted immediately after adding the solution with a Centro LIApcLB 692 plate luminometer (Berthold Technologies, Bad Wildbad, Germany). ROS evaluation was performed at a 2 min interval reading time for a period of 25 min.