Sample Contamination: Microbial, Chemical and Cross-Contamination

Contamination can ruin a sample silently. The peptide may look fine while microbes digest it, another compound distorts the results, or a leachable compound produces stray peaks. Each type has a different source and a different fix.

Microbial contamination

Peptides are a good nutrient source, and solutions near neutral pH at room or refrigerator temperature can support bacterial and fungal growth. Microbes also secrete proteases, which can cleave a peptide long before the solution looks cloudy.

• Use sterile water or solvent, sterile tubes and fresh tips.
• Prepare solutions in a clean area, ideally a biosafety cabinet for anything going into cell culture.
• Sterile-filter with a 0.2 µm low-protein-binding filter where the application requires it.
• Keep solutions cold and use them within your stated use-by period.
• Discard any solution that becomes cloudy or develops particles or odour.

Proteases from people

Skin carries proteases and microbes. Wear gloves, change them after touching surfaces outside the work area, and avoid touching tube rims, caps and tips.

Chemical contamination

• Leachables: plasticizers, slip agents and polymer additives can migrate from tubes and filters, especially with organic solvents. They often appear in mass spectra as repeating peaks.
• Detergent residues from washed glassware can suppress ionization in mass spectrometry and affect assays.
• Metal ions from spatulas, water or reagents can catalyze oxidation.
• Peroxides in aged ethers or some polymer surfactants oxidize Met and Cys.

Cross-contamination

Peptide powders are light and electrostatic. A spatula used for one peptide and wiped can still carry enough to show up in a sensitive mass spectrum of the next.

• Use disposable spatulas and weighing boats, or clean tools thoroughly between compounds.
• Open one compound at a time and close it before opening the next.
• Use separate, labeled solvent aliquots rather than a shared bottle.
• Use filter tips for pipetting stocks.

If you suspect contamination

1. Quarantine the tube and record what you observed.
2. Check other aliquots from the same preparation.
3. Test if the material is valuable: HPLC for new peaks, mass spectrometry for unexpected masses.
4. Review the preparation record to find the source, then discard affected material.