We need to test the effect of a peptide on gene expression in a tissue culture. We would like to make a stock solution but we don´t know which solvent to use? Does anyone have any experience with dissolving peptides for ex vivo culture? Suggestions in the literature include: phosphate buffer, DMSO, water, and media, but we don´t know which is the best for making a stock solution.
It depends on the composition of your peptide. DMSO works well for most, but it can do strange things to cells. If the peptide is not very hydrophobic, I personally prefer a slightly acidic solution (3-10mM HCl) to make a 1000X stock, and then dilute either in phosphate or medium.
Dear Carlos,
when I work with FPR receptor antagonist (BOC-2) that is a peptide. I diluted it in 50% MeOH.
Take a look at this link http://www.lifetein.com/handling_and_storage_of_synthetic_peptides.html#q2
it could help you.
Regards
If you know the peptide sequence, you can use this: http://www.innovagen.se/custom-peptide-synthesis/peptide-property-calculator/peptide-property-calculator.asp
handy tool to get some insights into your peptide properties.
I make 10 mM stock dilutions of peptides in 20% acetonitrile. You can also use these recommendations: http://www.sigmaaldrich.com/life-science/custom-oligos/custom-peptides/learning-center/peptide-solubility.html. Good luck!
If the peptide is soluble in aqueous solution, I would typically dissolve in TC grade PBS or 10mM HEPES at 1000X concentration and store the working aliquot at 4C. IF not, 5% DMSO is great as it doesn't freeze at 4C. Any left over peptide that you want to store long term should be lyophilized and stored at -20 to -80C. I've mass spec'ed peptides right after synthesis and after weeks or months of storage at 4C and even at -20C. Unfortunately there's quite a bit of degradation (like deamination or oxidation etc, depending on your sequence), so it's best to keep it in the powder form until you're ready to use it. Sometimes there's left over TCA or other buffers from synthesis, I like to remove these by dialysis or PD-10 columns before use in TC if you are using sensitive cell types.
Thank you very much to all
I have been able to dissolve my peptides with your recommendations. Quickly I explain to you: We have two peptides of 12 residuals each one. One of them has a negative overall charge and we have dissolved it in PBS without problems. The second one also has a negative overall charge but 50% of residuals are hydrophobic. For start we add PBS and the result has been that the peptide formed a gel solution. So we decide to add glacial acetic acid until a final concentration of 20%. The result was a perfect dissolution of the peptide. Being very concentrated it won't be a problem for their use in our ex vivo assays.
Thank you again to all.
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