ICE-based Mass Spectrometry Analysis

The freshly thawed samples were treated with deionized water or normal saline directly before the analysis. All samples were analyzed using the ICE technique and an LTQ Orbitrap XL ETD (ThermoFisher Scientific, San Jose, CA, USA) mass spectrometer as described elsewhere [14 (link)]. Briefly, the sample was placed inside a lab-made disposable cartridge (made from consumables provided by IDEX Health & Science LLC., Oak Harbor, WA, USA and GE Healthcare, Chicago, IL, USA) attached to the ESI interface of a mass spectrometer. The solvent flow (90% v/v methanol, 0.1% v/v acetic acid) extracts the analytes from the samples and delivers them into the ion source, while a glass microfiber filter inside the cartage protects the ESI needle from clogging. Mass spectra were acquired at the FTMS mass resolution set at 30,000 (FWHM at m/z 400 Da) for m/z values ranging from 500 to 1000 in the negative and positive modes, consecutively (30 s per polarity, 16 segments total). After the acquisition the ion source was washed with the solvent until the intensity of the signal decreased by at least 2 orders of magnitude from the level at the end of acquisition to avoid cross-contamination.