How does CM H2DCFDA work?
CM-H2DCFDA is a chloromethyl derivative of H2DCFDA, useful as an indicator for reactive oxygen species (ROS) in cells. This indicator exhibits much better retention in live cells than H2DCFDA. Subsequent oxidation yields a fluorescent adduct that is trapped inside the cell, thus facilitating long-term studies.
What is DCF fluorescence?
Dichlorofluorescein (DCF) is an organic dye of the fluorescein family, being substituted at the 2 and 7 positions by chloride. It is used as an indicator for argentometry by Fajans method. Dichlorofluorescin (DCFH) is a probe that is trapped within cells and is easily oxidized to fluorescent dichlorofluorescein (DCF).
What does DCFDA measure?
DCFDA / H2DCFDA / DCFH-DA / DCFH is a fluorogenic dye that measures hydroxyl, peroxyl and other reactive oxygen species (ROS) activity within the cell.
How do you dissolve H2DCFDA?
It can be dissolved in EtOH, DMSO and DMF at least 10 mg/ml. DMF is probably a better solvent for better stability. The stock solution is stable at least 6 months at –20°C. If H2DCFDA is prepared in aqueous buffer, dissolve before in 0.1M Na2CO3 and then in PBS to the desired concentration or pH.
How do you test for ROS?
Assess ROS by immediately analyzing your cells by flow cytometry using the FL1 channel (green fluorescence), or by fluorescence plate reader, or by fluorescence microscopy. The fluorescence can be detected by using excitation and emission wavelengths that are appropriate for green fluorescence.
What type of dye is fluorescein?
Fluorescein (lactone form) is a xanthene dye that is highly fluorescent, detectable even when present in minute quantities. Used forensically to detect traces of blood, in analytical chemistry as an indicator in silver nitrate titrations and in microscopy. It has a role as a radioopaque medium and a fluorescent dye.
What does flow cytometry tell us?
Flow cytometry provides a well-established method to identify cells in solution and is most commonly used for evaluating peripheral blood, bone marrow, and other body fluids. Flow cytometry studies are used to identify and quantify immune cells and characterize hematological malignancies.