CellROX™ Deep Red Reagent, for oxidative stress detection
CellROX™ Deep Red Reagent, for oxidative stress detection
Invitrogen™

CellROX™ Deep Red Reagent, for oxidative stress detection

CellROX™ Deep Red Reagent is a novel fluorogenic probe for measuring cellular oxidative stress in both live and fixed cellRead more
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Catalog number C10422
Price (USD)
626.00
Each
Add to cart
Price (USD)
626.00
Each
Add to cart

CellROX™ Deep Red Reagent is a novel fluorogenic probe for measuring cellular oxidative stress in both live and fixed cell imaging, with absorption/emission maxima at ∼644/665 nm. The cell-permeant dye is non-fluorescent while in a reduced state, and exhibits bright fluorescence upon oxidation by reactive oxygen species (ROS).

Features of the CellROX™ Deep Red Reagent:
• Optimized probe for detection of oxidative stress in cells
• Simple protocol that can be used with other live cell dyes and GFP
• Compatible with both live cell fluorescence-imaging and formaldehyde-based fixation methods

Substrate Specifics
The CellROX™ Deep Red reagent is a novel cell-permeant dye with absorption/emission maxima of ∼644/665 nm. CellROX™ Deep Red reagent is non-fluorescent while in a reduced state and becomes fluorescent upon oxidation by reactive oxygen species with emission maxima ∼665 nm that is measurable by fluorescent imaging, high content imaging, fluorescent plate readers, or flow cytometry.

Oxidative Stress Detection with CellROX™ Deep Red Reagent
Oxidative stress results from an imbalance between production of reactive oxygen species (ROS) and the ability of cells to scavenge them. ROS play an important role in the progression of several diseases including inflammation, atherosclerosis, aging, and age-related degenerative disorders. CellROX™ Deep Red Reagent can detect the oxidative stress in cells by reacting with ROS to become brightly fluorescent.

Simple, Robust Protocol
The protocol to use this reagent is simple and the bright deep red fluorescence signal is compatible with other live cell dyes and GFP, making it useful in multiplex fluorescence assays to measure a variety of cellular phenomena, including parameters related to cytotoxicity and cell death (Figure 4 below). Furthermore, unlike many other ROS sensors, such as H2DCFDA, the signal from CellROX™ Deep Red is retained after formaldehyde fixation for flexibility with the assay and improved workflows compared to those based on classic dyes for ROS detection.

For Research Use Only. Not for use in diagnostic procedures.
Specifications
ColorDeep Red
FormatLiquid
Quantity250 μL
For Use With (Equipment)Imaging, HCS, Cytometer
Concentration2.5 mM stabilized solution in DMSO
Detection MethodLive Cell Imaging
Excitation/Emission644/665 nm
IndicatorOxidative stress
Product LineCellROX™
Unit SizeEach
Contents & Storage
Store at ≤–20°C. Protect from light and desiccate.

Frequently asked questions (FAQs)

I want to assay cells for reactive oxygen species using carboxy-H2DCFDA, but I want to do so with a plate reader instead of microscope. Will it work?

It has been done. The problem is that plate readers are less sensitive than microscopes, with far less signal-to-background difference. It is worth trying, but first optimize concentrations and loading times with control cells, use a plate with little to no autofluorescence, and possibly optimize the gain setting in order to get the best signal possible. But don't expect the same sensitivity, even with optimization.

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

I have GFP-transfected cells and need to label for reactive oxygen species. Can I use H2DCFDA?

This is not recommended as the two dyes overlap in the emission wavelength. There are other ROS reagents available in different wavelengths, such as CellROX Deep Red, which emits in the far-red range (665 nm), or dihydroethidium, which is emits in the visible red range (620 nm).

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

I need a formaldehyde-fixable reactive oxygen species detection assay. Is H2 DCFDA fixable?

H2DCFDA and similar derivatives are not fixable. The same goes for dihydroethidium and dihydrorhodamine. However, CellROX Deep Red and CellROX Green are retained for a limited time upon fixation with formaldehyde. CellROX Green may be retained upon subsequent Triton X-100 permeabilization. Avoid the use of any acetone or alcohol-based fixatives or fixatives that include alcohol, such as formalin.

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

What dyes can I use to detect reactive oxygen species (ROS) in my bacteria?

Many dyes that are used on mammalian cells have also been shown to be useful in bacterial cells. For example, CellROX Deep Red Reagent has been shown to work in B. subtilis (see Reference: http://www-brs.ub.ruhr-uni-bochum.de/netahtml/HSS/Diss/RaatschenNadja/diss.pdf). If you are interested in a particular dye, but are not sure if it will work on your bacteria, literature searches are the best way to check to see if it has been tested. If not, then it may be worth testing yourself.

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

I am trying to label my cells with a reactive oxygen species (ROS) indicator dye, but I am not seeing a significant difference in signal. What could be happening?

First, make sure you have both a negative (untreated) and positive (ROS-induced) sample to compare. A good positive control can be the use of 100 µM menadione for one hour or 50 µM nefazodone for 24 hours. H2O2 can also be used, though it does not work well for CellROX dyes. Some dyes, such as H2DCFDA, require esterase cleavage, so don't incubate in the presence of serum (which contains esterases that can prematurely cleave the dye). If your positive control does not show significant change compared to the negative control, try increasing the concentration and label time for the dye. Our manuals give starting recommendations. Be sure to image your live cells as soon as possible. Only two dyes (CellROX Green and CellROX Deep Red) are retained with formaldehyde fixation. Finally, make sure you are using filters and instrument settings to match the excitation and emission spectra of the dye.

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.