Normally DMSO or DMF is recommended to dissolve DCF-DA ( here I am referring DCF-DA instead of DCF as you said in your question) and you can make small aliquotes and store at -20 C for several months. I hope that will help.

dissolve dch in dmso, presenve in small aliquots, covered to avoid any king of light and keep in at -80C°

Totally agree with the DMSO. After aiquote you can wrap the tube up with aluminum foil then put into -20C for at least weeks.

If you don't want to wrap every single tube in alumina, aliquot working volumes and wrap the box were you ar going to store the tubes

Thanks all of you. It seems that store under nitrogen or argon is not necessary... That solves most of my problem. Thanks!

I find i that the probe always degrades after making up if not stored properly. I find it best to make it up in DMF 10mg/ml and aliquot. Each aliquot gets gassed with N2 and placed into 50ml plastic centrifuge tubes, which are also gassed and sealed with para film and placed in a -35oC freezer. Aliquots are never exposed to light and used just before exprimentation in subdewed lighing. We have found that differing cell types require differing concs to get good results.

Dissolving DCF in DMSO is for solubility issues. Keeping it in the dark by wrapping in aluminum foil is to prevent photobleaching. Storing under nitrogen or argon is to prevent oxidation of the DCF which renders it useless as an indicator of oxide production in your experiments. If you don't have nitrogen or argon available you will have to determine empirically how long the DCF remains usable after being stored in frozen aliquots without the nitrogen or argon. My bet would be a few months, but that's just a guess. I don't know for sure.

We usually order 20x50ug package (http://products.invitrogen.com/ivgn/product/C6827). You can keep unopened ones for at least a year in minus 20. One 50ug tube should be enough for one set of experiment, depending on your conditions.

We do not dissolve it, but store as dry powder and make fresh solutions the day we need it.

This us why DCF is not a good probe: unstable and auto-oxidizes. I prefer dihydro-rhodamine [detects all ROS] or APF [OH radical].

For sure dihydro-rhodamine is better, but all of them have some limitations. For example, APF is also oxidized by peroxinitrite.

I recomend you to storage severals alicuots (20 ul) of it in DMSO at -20°C, and then cover each micro-tube with aluminum foil.

Good luck!

dissolve it in DMSO and store at-20C in dark bottle. prefer to make 20-50 ul of aliquots to prevent freeze thaw

Storing at -20 after dissolving in DMSO after wrapping in al foil is the best. Storage life best between 2weeks to 1 month that is what I felt best.

Our experience indicates that the solution aliquoted and stored at -20 ° C is stable for several months

Remember that DMSO has mild reducing properties. This should protect most solutes from oxidation during storage.

Do you refer to DCF or DCF-DA?? I usually use the DCF-DA (2',7'-dichlorofluorescein diacetate) because it is more stable than DCF since the two reactive OH-groups are protected from oxidation by acetate groups through ester bonds. This bond is stable and can be broken only inside the cells by esterase enzymes so the molecule can passively diffuse into cells and is colorless and nonfluorescent until the acetate groups are cleaved by intracellular esterases to yield the fluorescent fluorophore.

Like DCF,it is soluble in DMF, ethanol (2 mg/mL), DMSO (10 mg/mL), can be stored at -20°C for up to six months but my experience indicates that you can also use it after 6months.

You may want to aliquot in DMSO because of solubility. Wrap with foil paper and minimal exposure to light when handling is vital. If stored at -20 or -80 oC may preserve the integrity for more than 3 months.

I agree with Barbara. We also use DCF-DA, it is quite stable, just keep it protected from light. We store it at -20C, and had no problem in longer storage.

Diacetate is removed by esterases, and then the dye can be oxidazed inside the cell.

I recommend you to storage severals aliquots (20 ul) of it in DMSO at -20°C. I keep it protected from light. And I store it at -20C, and have no problem in longer storage.

In my practice, solubilization in pure DMSO, in minimum volume u can use and store at -20 is good. I have checked the stability of certain oxidizable materials are stable for more then 6 months. Every time use less quantity of aliquots to preserve and foil. If u can use amber colored or any dark tube for aliquots ,its preferable. Allow single use of aliquot and minimal exposure to light.

For DCF-DA, by dissolving in methanol (1.25mM) as small aliquots in eppendorf and wrapped by aluminium foil and kept in -20 is good to store for a month

Followed by previous comments, I agree, just dissolve in DMSO. Then make numerous aliquots and keep them frozen in -20C. Protect from light !!!

I agree with my colleagues, but I dissolve 2,7dichlorofluorescein diacetate in ethanol and store in room temp protected from light during 1 month.

You have no problem, when you work with 2,7 dichlorofluorescin diacetate.

We handle the dissolved ester as Marcos Pereira recomended to you.

Thanks all of you again. I have titre my DCF-DA with the protocol recommended by you: just to dissolve it in DMSO. The concentration is 20mM, so that I only have to add 1/1000 DCF-DA into the medium.

By the way, I have almost completed my experiment, the result is good, even beyond my expectation:)

Thanks again.

the recommendation of provider (http://www.enzolifesciences.com/ALX-610-022/2-7-dichlorodihydrofluorescein-diacetate/)

Stock solution in organic solvents is stable for at least 6 months if stored at -20°C. For maximum solubility in aqueous buffers, dissolve directly in 0.1M disodium carbonate (7.7mg/ml) and then immediately diluted with PBS, pH 7.2, to the desired concentration or pH. Upon neutralization of the solution the concentration may not be greater than 1mg/ml. We do not recommend storing the aqueous solution for more than one day.

We store the solutions under nitrogen. Not clear whether low temperature is needed, since this will freeze the DMSO and concentrate the drug in the last bit to freeze. This will not occur with ethanol. DCF is not the ideal drug since it does tend to autooxidize and measures multiple reactive oxygen species. For OH radical, APF is best. For singlet oxygen, DADB. Since singlet oxygen is rarely formed, check the specificity of APF and see if this isn't satisfactory for what you are doing.

DMSO does not crystallize like water does. It forms a continuous phase that is oft and malleable. Solutes remain dispersed after freezing, which is one reason DMSO is the standard storage medium for drugs.