ROS-Glo™ H2O2 Assay

ROS assay cell type - PANC-, BxPC-3 human pancreas

Experiment
ROS assay cell type - PANC-, BxPC-3 human pancreas
Product
ROS-Glo™ H2O2 Assay from Promega
Manufacturer
Promega

Protocol tips

Protocol tips
- H2O2 substrate was added together with SW43-DOX.

- After 6h, detection solution was added and luminescence was measured 20 min.

Publication protocol

Caspase 3/7 activity was measured using the Caspase-Glo® 3/7 Assay (Promega, Madison, WI, USA). Briefly, the assay is based on a caspase 3/7 substrate resulting in a caspase-specific luminescence signal equivalent to the caspase 3/7 activity. ROS Quantification was performed using the ROS Glo™ Kit (Promega, Madison, WI, USA). Here, the H2O2 substrate reacts with H2O2 generating a Luciferin precursor which is converted to light emitting Luciferin through a detection solution. The luminescence signal is proportional to the H2O2 concentration. Panc-1 cells were treated with 25–50 μM SW43-DOX for 6 h. Pan-caspase blocking (25 μM Z-VAD-FMK; R&D Systems, Minneapolis, MN, USA) or ROS elimination (200 μg/ml α-Tocopherol) was started 1 h prior to treatment. Caspase 3/7 substrate was added onto cells after treatment medium removal and luminescence was measured 30 min thereafter. H2O2 substrate was added together with SW43-DOX. Six hours later, the detection solution was added and luminescence was measured 20 min thereafter. Luminescence was quantified using the multi-mode microplate reader. Caspase 3/7 activity was normalized to parallel measured viability to account for occurred cell death by treatment and by untreated control group. Each sample was tested in at least triplicates.

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Papers

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Paper title
[Bibliography of communications and reports by Hungarian authors, published outside the Fogorvosi Szemle].
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Manufacturer protocol

Download the product protocol from Promega for ROS-Glo™ H2O2 Assay below.

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