Leaf discs (0.196 cm2 (link)) of 4-week-old WT plants were incubated overnight floating on 0.1 mL water in 96-well titer plate, with one disc per well. The following day the leaf discs were pre-treated with 100 nM rAtPNP-A or mock for 30 min. For ROS detection horseradish peroxidase and luminol (Sigma-Aldrich) were added to a final concentration of 10 μg mL−1 and 100 μM, respectively. Luminescence was measured directly after addition of either 1 μM of flg22 or 100 nM rAtPNP-A in a MicroLumat LB96P plate reader (Berthold Technologies) for 1 h and is shown in relative light units.
Microlumat lb96p plate reader
Manufactured by Berthold Technologies
Sourced in Germany
The MicroLumat LB96P plate reader is a compact and versatile instrument designed for luminescence-based assays. It features a 96-well plate format and is capable of performing sensitive measurements of various luminescent signals.
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Lab products found in correlation
2 protocols using microlumat lb96p plate reader
1
Luminescence-based ROS detection assay
2
Measuring HIF-1 Activity in Cell Lines
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MCF-7, Hep3B and U2OS cells were used to measure HIF-1 activity in reporter gene studies. Cells were allowed to grow on 24-well plates before they were transiently transfected with a hypoxia-responsive luciferase plasmid which contains six HIF-1 binding sites from the transferrin 3′ enhancer.37 (link) Empty control vector (pGL4 vector, Promega, Mannheim, Germany) served as transfection control. Medium was renewed 24 hrs after transient transfection. Cells were pre-treated with 0.1 or 1.0 µM selinexor or DMSO for 1 hr before incubation in normoxia (20% O2) or hypoxia (1% O2) for 24 hrs. After incubation, lysis was performed using Reporter Lysis 5X Buffer (Promega). Luminescent signal intensities were analyzed with the Luciferase Assay System (Promega) using a MicroLumat LB 96P Plate Reader (Berthold Technologies, Bad Wildbad, Germany). Bio-Rad DC Protein Assay (Bio-Rad, München, Germany) was used for determination of protein concentrations. Firefly luciferase (FL) activities were normalized to protein concentrations. Three technical replicates were analyzed.
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