I was trying to dissolve PMSF in lysis buffer. But due to little water solubility it couldn't get dissolve. Which solvent I should use to dissolve it? I will use this lysis buffer for the protein extraction for WB. I am wondering if ethanol or isopropyl alcohol will interfere with protein extraction or their stability. Please suggest. Thanks chandrashekhar.
I make a stock solution of 100 mM PMSF in isopropyl alcohol, then dilute it into the needed buffer at the last minute because it is unstable especially at high pH. This circular from Sigma Aldrich shows how long PMSF is effective in aqueous solutions of different pH. https://www.sigmaaldrich.com/content/dam/sigma-aldrich/docs/Sigma/Product_Information_Sheet/1/p7626pis.pdf.
Another reagent, AEBSF that has inhibitor action similar to PMSF, is soluble in aqueous solution. Here is the product information. https://www.sigmaaldrich.com/content/dam/sigma-aldrich/docs/Sigma-Aldrich/Product_Information_Sheet/a8456pis.pdf
Hi there,
0.1M stock (X100) in absolute ethanol stored at -20°C is OK. Add to cold lysis buffer right before use. 1% ethanol in lysis buffer has no effect on protein.
Hi Chandrashekhar, you can also make a 100-1000x stock solution of PMSF in anhydrous DMSO.
Hi, make a stock of 100mM PMSF in 100% ethanol or isopropyl alcohol and use a final concentration of 0.1mM in the lysis buffer. the organic solvent at this concentration has no effect on the proteins.
I make a stock solution of 100 mM PMSF in isopropyl alcohol, then dilute it into the needed buffer at the last minute because it is unstable especially at high pH. This circular from Sigma Aldrich shows how long PMSF is effective in aqueous solutions of different pH. https://www.sigmaaldrich.com/content/dam/sigma-aldrich/docs/Sigma/Product_Information_Sheet/1/p7626pis.pdf.
Another reagent, AEBSF that has inhibitor action similar to PMSF, is soluble in aqueous solution. Here is the product information. https://www.sigmaaldrich.com/content/dam/sigma-aldrich/docs/Sigma-Aldrich/Product_Information_Sheet/a8456pis.pdf
Hi Chandrashekhar
you can solve it in isopropanol or ethanol also. because pmsf is instable don't add it in your whole lysis buffer stock. you should make it just before you want to add it in your lysis buffer.so it is better to prepare a 200 or 100 mM stock of pmsf and then use it in 0.1-1mM in final volume of your lysis buffer .
Will adding stock solution of PMSF to TE buffer (pH 8) inactivate PMSF , as it is unstable in water.
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