DCFDA / H2DCFDA is a cell permeable fluorescent probe that is redox sensitive and used to measure the concentration of reactive oxygen species (ROS) within a population of cells. DCFDA / H2DCFDA diffuses into cells where it is hydrolysed by intracellular esterases into a non-fluorescent and non-cell permeable intermediate. Upon reaction with ROS this forms the fluorescent compound 2’,7’ –dichlorofluorescein (DCF) which is excited at 485nm and emits at 535nm. Pyocyanin is included within this kit as a positive control. Pyocyanin promotes the formation of ROS through inactivation of catalase and depleting reduced glutathione.
This kit contains:
DCFDA / H2DCFDA assay reagent
DMSO
Lyophilised Pyocyanin
10x assay buffer
Description
Kit for measurement of reactive oxygen species (ROS) within cells.
Figure 1. Concentration response of pyocyanin in HEK293T cells using HB7375 DCFDA / H2DCFDA - Cellular ROS Assay Kit
Pyocyanin induces oxidative stress in HEK293T cells which is detected as an increase in fluorescence using the H2DCFDA probe. Method: HEK293T cells were seeded at 50,000 cells per well in a 96 well microplate before being cultured overnight. The following day cells were washed with 1x assay buffer before being incubated in 20µM H2DCFDA for 45 minutes at 37°C. Cells were then washed again with 1x assay buffer before being incubated in differing concentrations of pyocyanin in assay buffer for 1 hour at 37°C. Fluorescence was then read in a microplate reader (excitation at 485nm and emission at 535nm).
Figure 1. Concentration response of pyocyanin in HEK293T cells using HB7375 DCFDA / H2DCFDA - Cellular ROS Assay Kit
Pyocyanin induces oxidative stress in HEK293T cells which is detected as an increase in fluorescence using the H2DCFDA probe. Method: HEK293T cells were seeded at 50,000 cells per well in a 96 well microplate before being cultured overnight. The following day cells were washed with 1x assay buffer before being incubated in 20µM H2DCFDA for 45 minutes at 37°C. Cells were then washed again with 1x assay buffer before being incubated in differing concentrations of pyocyanin in assay buffer for 1 hour at 37°C. Fluorescence was then read in a microplate reader (excitation at 485nm and emission at 535nm).
Optical Data
Emission color
Green
Max excitation wavelength
485nm
Max emission wavelength
535nm
Closest laser lines
488 nm
Cell permeable
Yes
Biological Data
Application notes
Please follow this link to a full DCFDA / H2DCFDA - Cellular ROS Assay Kit protocol
Solubility & Handling
Storage instructions
-20°C
Important
This product is for RESEARCH USE ONLY and is not intended for therapeutic or diagnostic use. Not for human or veterinary use