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FCS and Protein assay


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3 replies to this topic

#1 jakatta70

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Posted 02 June 2009 - 08:28 AM

Hi all,

I am using drugs dissolved in DMSO in Caco-2 FCS-free media but I have been told that my drugs are coming out of solution because I have not added any FCS. However I have avoided using serum media because I need to quantify proteins in the cell supernatant using Bradford assay. Is there a way I can account for the FCS in the media when measuring supernatant protein? I am using 5% FCS.

#2 bob1

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Posted 02 June 2009 - 04:42 PM

5% FCS will very likely swamp out you being able to detect any secreted proteins. I would be surprised if your drugs are coming out of solution because there is no FCS, more likely is that they are not very soluble in water.

#3 jakatta70

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Posted 03 June 2009 - 01:57 AM

Hi bob1,

Thanks for your reply. Yes the drugs I am using are very hydrophobic and polar with lots of benzene rings. They readily dissolve in 100% DMSO (which is what I have dissolved them in). I was just told yesterday by a former PhD student that she got the drugs to stay in solution by adding a small % of FCS plus warming the drugs at 37C before transferring to the tissue culture plates with media. I thought the former was a little strange but at this stage I am willing to try anything! I am also going to reduce the concentration of drugs from 50 to 30 uM but I doubt this will have a huge effect on solubility!

#4 bob1

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Posted 03 June 2009 - 04:11 PM

Try dissolving them in dimethyl fomamide, it may mean that they stay in solution better.




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