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Peptides were eluted over a 120-min gradient with buffer A as 100% water + 0.1% formic acid and buffer B as water + 0.1% formic acid and acetonitrile with the following steps: 05% B for 1 min, 525% for 68 min, 2540% for 20 min, 4060% for 15 min, 6095% for 1 min and 95% for 15 min
Notably, the affinity of cysteine for cysteinyl-tRNA synthetase is orders of magnitude higher than its affinity for the rate-limiting enzyme in GSH synthesis (GCLC) 48,49
The process is straightforward: we start with a simple blood draw, just like youd have at a doctors office
To confirm the MTT results, a replicate plate run in parallel was developed by adding 5 l of 2.5 M Calcein-AM (ThermoFisher Scientific) to selected wells and incubated 15 min at room temperature
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