The best way to handle solubility problems, is to keep the DMSO concentration as high as possible in the final assay, and dilute directly from your DMSO stock into the cells with media. Also it is critical to make the serial dilutions in DMSO if you are doing a dose response. So let's say the cells can take 0. 1-0.2% DMSO as a final concentration. Then if you have 1 ml volume, you can add 1-2ul of your compound directly into the 1 ml.

I don't remember what the maximum concentration your cells can tolerate, but I would use that concentration and work backwards to determine how many ul you need to add. This is really the only way to deal with insoluble compounds.

So if your final concentration is 1 uM, then I would add 1ul from a stock of 500uM in DMSO only, to 500ul of media with cells. Or you can try to make a 10% solution of DMSO with a 50 uM stock, and add 5ul. If you make the 10% stock, it is risky, but you can see if it's precipitates. If your cells can tolerate more DMSO than 0.1-0.2%, then you can add more ul of a more dilute stock solution.

Hi Monika,

I've experienced this exact same problem before, but found with subsequent dilutions the final concentration was clear. In other words, I would always make up my initial stock in DMSO around 1000-10,000 fold more concentrated than required. For example you might dissolve 1 mg of reagent in 50-100 microlitres of DMSO (which should be clear), then serially dilute your sample as required with good mixing in between until you reach your final desired working concentration. As the concentration of your material becomes less, the degree of solubility in culture medium or other physiological solution should increase to level where there is no precipitate, but of course this will vary between different materials. What are you working with?

Yes, I did exactly this. I dissolved my compound in 50ul of DMSO (100mM). And for further dilution I took 0.6ul of DMSO solution in 999.4ul of water to get 100uM. It looks like that I dissolved, But, I still can see particles into it . I have to take 5ul of this 100uM into 500ul of media (1uM). But, if particles remains I can not say that it is 1uM concentration.

Moreover, I want to filter it prior to use in cell culture, but if particles remain , then probably, filteration will not be a good option.

I am treating BLT-1 inhibitor.

OK, sounds like you're on the right track.

Perhaps particles are impurities?

I understand your concern though regarding actual concentration at end.

Would suggest filtering and then testing to see if you get expected level of activity.

Perhaps also re-check any publications where they have done similar and check with chemical supplier for any advice (product information sheets etc).

Good luck!

Yes, thank you.

The best way to handle solubility problems, is to keep the DMSO concentration as high as possible in the final assay, and dilute directly from your DMSO stock into the cells with media. Also it is critical to make the serial dilutions in DMSO if you are doing a dose response. So let's say the cells can take 0. 1-0.2% DMSO as a final concentration. Then if you have 1 ml volume, you can add 1-2ul of your compound directly into the 1 ml.

I don't remember what the maximum concentration your cells can tolerate, but I would use that concentration and work backwards to determine how many ul you need to add. This is really the only way to deal with insoluble compounds.

So if your final concentration is 1 uM, then I would add 1ul from a stock of 500uM in DMSO only, to 500ul of media with cells. Or you can try to make a 10% solution of DMSO with a 50 uM stock, and add 5ul. If you make the 10% stock, it is risky, but you can see if it's precipitates. If your cells can tolerate more DMSO than 0.1-0.2%, then you can add more ul of a more dilute stock solution.

Check researchgate answers to tolerability of DMSO by cells. Google below Question and see Researchgate answers. 0.1% is typical! but some cells can go higher. hepatocytes can go to 1%. To be safe, I would use 0.1% .

Question

Does anybody know what is the safe solution of DMSO for cell cultures?

Im naive to this field,

but still as im currently working i would like to suggest

U dissolve your sample in higher concentration so dat u endup adding max.0.1%DMSO solution to you cells.

at this concentrations DMSO is not toxic to cells.

hope this will help u.

ALL THE BEST

Thanks all. I will try with putting DMSO directly into the cells and will see.

Thanks

When you do that, put the same amount of DMSO (no drug) in a control flask, as control for vehicle.

Yes, Ana I do put control always.

Thanks

Thanks Elizabeth, I was thinking of doing something like this.

Hi

I confirm that in the most of cases, there is not cell toxicity by using DMSO in a final dilution of 1/1000. In order to facilitate DMSO dissolution , you can try to heat your medium before the addition of DMSO. DMSO is very temperature sensitive.

Best regards

Olivier

I generally try to avoid using any DMSO in cell culture methods. While DMSO may not be toxic it definitely indices many significant changes in gene expression and these can complicate interpretation of the observed changes in gene expression that we are looking for.

Hi again,

DMF is another option from memory, but I like the suggestions to pre spike the medium with 0.1% DMSO and try heat if required.

I already had this problem, Absolute ethanol (Merck) replace it. of course, I don't know about your material and solution which you able to use it,

DMSO may affect on gene expression, you will earn best result, if you use final dilution what Olivier Delezay said

If you are doing it for cell culture, dilute it into serum-containing medium. If necessary, you can add it initially to a very high concentration of albumin or serum and then dilute into tissue culture medium. You want the compound to be adsorbed to serum proteins.

@ : Jan kola: is that the advertisement in Researchgate.net ?? :)

Dissolve your substance in the minimum amount of DMSO(a few microlitres). Then dilute with an appropriate buffer to the required concentration.

Generally solubility of pure compounds may differ, based on the manufacturer instructions we can choose the solvents. 0.1  to 0.5% of  DMSO is not toxic to cells at the same time we can dissolve the compounds in higher concentration of  DMSO then only compound completely dissolved. But further dilute in to very minimum concentration based on pipetting  accuracy for assays. sodium salts etc.. containing compounds only soluble in water or saline and rest of the pure compounds are only soluble in solvents like methanol, ethanol and DMSO etc....

Hello Monika, I am facing a similar problem with my chemical. Did all the above suggestions help with the precipitation problems of yours?