Invitrogen™ CellROX™ Reagent for oxidative stress detection

Product Code.	Color	Unit Size
12013765	Deep Red	Pack of 5
12887552	Orange, Far-Red, Green	Each
12648335	Green	250µL
12807322	Orange	50µL

Product Code. 12013765 Supplier Invitrogen™ Supplier No. C10422

Description

Includes

5 x 50 μL, 2.5 mM stabilized solution in DMSO

CellROX™ Reagents are novel fluorogenic probes for measuring cellular oxidative stress in both live and fixed cell imaging.

The CellROX™ Reagents are novel fluorogenic probes for measuring cellular oxidative stress in live cells. These cell-permeant dyes exhibit bright fluorescence upon oxidation by reactive oxygen species (ROS). (See table below for details of each reagent.)

The CellROX™ reagents are:

• Optimized for detection of oxidative stress in live cells and may be added directly to serum-containing growth media
• Provided as a stable, ready-to-use DMSO solutions with simple protocols, compatible with standard workflows in fluorescence microscopy
• Compatible with a variety of detection platforms, including traditional fluorescence microscopy, high content screening (HCS), and flow cytometry

Oxidative stress results from an imbalance between the 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.

For Research Use Only. Not intended for any animal or human therapeutic or diagnostic use.

Order Info

Shipping Condition: Wet Ice

Specifications

• Color: Deep Red
• Format: Liquid
• Quantity: 5 x 50 μL
• Content And Storage: Store at ≤–20°C. Protect from light and desiccate.
  
  
• For Use With (Equipment): Imaging, HCS, Cytometer
• Concentration: 2.5 mM stabilized solution in DMSO
• Detection Method: Live Cell Imaging
• Excitation/Emission: 644/665 nm
• Indicator: Oxidative stress
• Product Line: CellROX

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.

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).

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.

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.

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.

For Research Use Only. Not for use in diagnostic procedures.