E coli sample prep for LCMS

Question

This question concerns sample preparation for LCMS. I am preparing an extract of peptides from Ecoli (ATCC 25922) for identification by LCMS per the following reference: PMID: 26510657 , J. Am. Soc. Mass Spectrom. (2016) 27:194-210. After cell growth, the sample is centrifuged, washed, and then resuspended in 70% formic acid. Due to the non-volatility of formic acid, we are having difficulty drying the sample. We don't have a lyophilizer, so do you have any simple ideas we could try to get us through this point? We can make any modifications to the prep to achieve the desired result of extracted peptides. Best regards.

andyg · Accepted Answer

You will want to look for a solvent system that dissolves the pellet.

Maybe try 50% ACN with a lower amount of acid (maybe 0.1% formic acid or 0.2% acetic acid). 70% HFIP (hexafluoro-isopropanol) might work well too. Either solvent system may work well without the addition of acid.

The next step after centrifugation is the addition of ACN. You will want to keep the organic concentration low enough to prevent protein precipitcation but high enough to help solubilze the proteins- maybe 50-70% organic