Peptide Probe-Based NanoLC-MS/MS PRM for Ultrasensitive Detection of SARS-CoV-2 Proteins in Environmental Samples

Objective This study aimed to develop a sensitive and reliable method for detecting severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in various environmental samples using nano–liquid chromatography–tandem mass spectrometry parallel reaction monitoring (nanoLC-MS/MS PRM), targeting sequences of the viral spike (S) and nucleocapsid (N) proteins. Materials and Methods A synthetic peptide screening approach was employed to systematically evaluate candidate sequences as surrogates for protein quantification. The process began with the digestion of purified SARS-CoV-2 proteins. The candidate sequences were then refined to a subset of target peptides, and stable isotope-labeled heavy peptides were used as standards in a PRM workflow. The detection limits were determined using the nanoLC-MS/MS PRM method. Results The developed method achieved limits of detection in the 1 fmol/µL (2–8 × 10-15 viral copies/L) range on-column. This demonstrates high sensitivity for detecting SARS-CoV-2 in environmental samples using the PRM method. Conclusion The study successfully developed a sensitive and reliable peptide probe-based approach for detecting SARS-CoV-2 in various environmental samples. The use of the PRM method targeting the S and N proteins of SARS-CoV-2 provides a robust approach for virus detection, particularly given the potential persistence of the virus on surfaces. Graphic Graphic . Open in a new tab