TFA is one of the acids that can be used for acidifying biological samples and is directly compatible with μPC-microLC-MS. We noticed, however, that TFA
While precipitation is an obvious choice for concentrating DNA and RNA samples, it can also be an effective way to concentrate proteins. I have tried some LC-MS-friendly detergent but they don't make much difference. Quantitative protein precipitation using TCA is specific to deoxycholate and certain other detergents in solution; TCA can be used alone for precipitation, but the pellet is not readily dissolved, causing sample losses [59,145]. This is critical and why this acid is put to this use. The basic answer is yes the peptides are denatured a bit with TFA, however 0.15 does not degrade the peptide. Protein precipitation can be performed by acidifying the sample or by adding an organic solvent, e.g.
In such cases, repeated precipitation may be performed. TFA is used in peptide HPLC as an ion pairing reagent to help separate the peptides in a reverse-phase chromatographic (HPLC) run.
• A single precipitation may not be sufficient to remove all types and concentrations of interfering contaminants. Are you using it with whole proteins or peptides? optional: (0.1% b-mercaptoethanol or 20 mM dithiothreitol) Equilibrate (0 oC) for 10-20 min. Here in installment two of this three part series, I describe the two most common methods for protein precipitation – ammonium sulfate and trichloroacetic acid. Trifluoroacetic acid (TFA) is an organofluorine compound with the chemical formula CF 3 CO 2 H. It is a structural analogue of acetic acid with all three of the acetyl group’s hydrogen atoms replaced by fluorine atoms and is a colorless liquid with a vinegar like odor. For precipitation before performing a BCA Protein Assay, see the Tech Tip #8: Eliminate interfering substances from samples for BCA Protein Assay. acetonitrile.
In a typical TCA-deoxycholate (DOC) precipitation, the protein solution is mixed with a dilute sodium deoxycholate buffer. I have done acetone precipitation, but it is very difficult to solubilize the protein pellet even with 8M urea.