Q: If I already removed small contaminants and detergents at the protein level, do I still need to desalt my peptides prior to mass spectrometry analysis?

Yes. We recommend performing additional cleanup after protein digestion to remove any residual salts or partially digested proteins using Pierce Peptide Desalting Spin Columns (Cat. Nos. 89851, 89852), Pierce C18 Spin Tips (Cat. No. 84850) or an in-line C18 trap column (https://www.thermofisher.com/order/catalog/product/164564-CMD). Find additional tips, troubleshooting help, and resources within our Mass Spectrometry Support Center .

Question 1

Do you offer the adaptors provided with Pierce C18 Spin Tips (Cat. No. 84850) as a stand-alone product?

Accepted Answer

Unfortunately, the adaptors provided with Pierce C18 Spin Tips are not sold separately and they are supplied in a smaller quantity because they are meant to be reused.

Find additional tips, troubleshooting help, and resources within our Mass Spectrometry Support Center.

Question 2

Do I need to do C18 clean-up of my mass spectrometry sample before phosphopeptide enrichment?

Accepted Answer

Yes. C18 clean-up is necessary to remove small molecules that may interfere with phosphopeptide enrichment. Drying after clean-up is also necessary to concentrate the dilute samples and remove C18 organic eluting solvent so samples can be dissolved in the optimized buffers for phosphopeptide binding. Alternative binding buffers are not recommended as phosphopeptide yield and specificity will be affected. For maximal performance, ensure that the clean, dried peptide samples are completely solubilized in binding buffer with agitation for at least 10 min.

Find additional tips, troubleshooting help, and resources within our Mass Spectrometry Support Center.

Question 3

How do I determine if my sample is compatible and ready for LC-MS?

Accepted Answer

Samples prepared using LC/MS grade reagents are suitable for LC-MS; however, particulates and other small molecules can all interfere with liquid chromatography separation and mass spectrometer source ionization. We recommend visual inspection of samples for particulate matter and using Pierce Peptide Desalting Spin Columns (Cat. Nos. 89851, 89852), Pierce C18 Spin Tips (Cat. No. 84850) or an in-line C18 trap column (https://www.thermofisher.com/order/catalog/product/164564-CMD) to remove non-volatile salts before MS analysis.

Find additional tips, troubleshooting help, and resources within our Mass Spectrometry Support Center.

Question 4

I cleaned up my peptide sample using C18 or desalting columns, but I lost most of my peptides. What went wrong?

Accepted Answer

Peptides do not bind well to reversed phase resins at neutral pH or in the presence of organic solvents (e.g., acetonitrile). Acidify protein digest samples using formic acid or trifluoroacetic acid (TFA) to pH <3 before desalting. Ensure that samples do not contain organic solvents before and after clean-up by drying them down using a SpeedVac concentrator or equivalent.

Find additional tips, troubleshooting help, and resources within our Mass Spectrometry Support Center.

Question 5

If I already removed small contaminants and detergents at the protein level, do I still need to desalt my peptides prior to mass spectrometry analysis?

Accepted Answer

Yes. We recommend performing additional cleanup after protein digestion to remove any residual salts or partially digested proteins using Pierce Peptide Desalting Spin Columns (Cat. Nos. 89851, 89852), Pierce C18 Spin Tips (Cat. No. 84850) or an in-line C18 trap column (https://www.thermofisher.com/order/catalog/product/164564-CMD).

Find additional tips, troubleshooting help, and resources within our Mass Spectrometry Support Center.

Pierce™ C18 Spin Tips, 96 Tips - FAQs

Do you offer the adaptors provided with Pierce C18 Spin Tips (Cat. No. 84850) as a stand-alone product?

Do I need to do C18 clean-up of my mass spectrometry sample before phosphopeptide enrichment?

How do I determine if my sample is compatible and ready for LC-MS?

I cleaned up my peptide sample using C18 or desalting columns, but I lost most of my peptides. What went wrong?

If I already removed small contaminants and detergents at the protein level, do I still need to desalt my peptides prior to mass spectrometry analysis?