Table 2 Conditions for turbulent flow chromatography (TFC) loading and eluting pumps for analysis of SARS-CoV-2 nucleoprotein target peptides.

Step	Start (min)	Loading pump							Eluting pump
Step	Start (min)	Flow	Grad	%A	%B	%C	%D	Tee	Loop	Flow	Grad	%A	%B
1	0	1.2	Step	100	‒	‒	‒	====	out	0.3	Step	99	1
2	0.5	1.2	Step	100	‒	‒	‒	====	out	0.3	Step	99	1
3	1	0.2	Step	100	‒	‒	‒	====	out	0.3	Step	99	1
4	1.1	0.1	Step	100	‒	‒	‒	T	in	0.3	Step	99	1
5	2.7	1.2	Step	60	40	‒	‒	====	in	0.3	Step	80	20
6	3.9	1.2	Step	‒	‒	‒	100	====	out	0.3	Ramp	75.5	24.5
7	4.9	1.2	Step	‒	‒	100	‒	====	out	0.3	Ramp	75.0	25
8	5.9	1.2	Step	‒	‒	‒	100	====	out	0.3	Ramp	72.5	27.5
9	6.9	1.2	Step	‒	‒	100	‒	====	out	0.3	Ramp	70	30
10	8.4	0.5	Step	100	‒	‒	‒	====	out	0.3	Ramp	50	50
11	8.6	0.5	Step	100	‒	‒	‒	====	out	0.3	Step	20	80
12	9.6	1.2	Step	100	‒	‒	‒	====	out	0.3	Step	99	1

Steps 1 to 3: sample loading; step 4: transfer of peptides onto the analytical column; steps 5 to 10: switching wash between organic solvents (C and D) to clean large particles and reduce carryover in the TFC column, and peptide elution from the analytical column; step 11: TFC column equilibration and analytical column washing; step 12: system equilibration.
Loading: A (1‒4) 0.5% formic acid in water; B (1‒4) acetonitrile; C (1‒4) isopropanol, acetonitrile, and acetone (40/40/20, v/v/v); D (1–4) 20% dimethyl sulfoxide (DMSO) and 2% trifluoroethanol in water.
Eluting: A (1‒4) 1% DMSO, 0.1% formic acid in water; B (1‒4) 1% DMSO, 0.1% formic acid in acetonitrile.

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