Uplc 1 class ftn system

LC separation of nucleotides (cAMP, AMP, ATP, cGMP, GMP, GDP, and GTP) was performed with a UPLC I-Class FTN system (Waters) equipped with an Atlantis Premier BEH C18 AX column (2.1 mm x 50 mm, 1.7μm, Waters). Separation was performed at a flow rate of 400 μl/min with the following LC gradient: 0–1 min: 3% B, 1–4 min: 3–20% B, 4–5 min: 20–85% B, and 5–5.5 min: 85% B; solvent A, 0.2% (v/v) formic acid in water; solvent B, 0.2% (v/v) formic acid in acetonitrile. The UPLC system was directly coupled to a Xevo TQ-XS mass spectrometer (Waters) with an electrospray ionization (ESI) source. Multiple-reaction monitoring (MRM) was performed using specific transitions for the nucleotides cAMP, AMP, ATP, cGMP, GMP, GDP, and GTP.

Four selected synthetic peptides derived from SARS-CoV-2 nucleoprotein (Arg41-Lsy61, Arg41-Lys65, Met210-Arg226, and Met210-Lys233) and their heavy isotope (¹³C and ¹⁵N)-labeled variants (Arg41-Lsy*61, Arg41-Lys*65, Met210-Arg*226, and Met210-Lys*233, SpikeTide-TQL peptides, where * denotes [¹³C₆¹⁵N₄]-Arg or [¹³C₆¹⁵N₂]-Lys) were purchased from JPT Peptide Technologies and used for method development and as quantitation standards. LC separation of peptides was performed on a UPLC I-Class FTN system (Waters) equipped with a BEH C18 column (2.1 mm × 50 mm, 1.7 μm, Waters). The UPLC system was directly coupled to a Xevo TQ-XS mass spectrometer (Waters) equipped with electrospray ionization (ESI) source. MS acquisition was performed using an MRM method of two selected transitions per peptide with optimized collision energies.