Leukocyte vitamin C measurements

The buffy coat layer was transferred to 2 mL cryovials and stored at −40 °C until analysis. A 500 µL aliquot was taken and 500 µL of 10 % PCA with 1 % metaphosphoric acid (MPA) was added. The mixture was vortexed in foil for 1 min. 500 µL of mobile phase was added and the mixture was vortexed for another minute and centrifuged for 10 min at 12,000 RPM. 20 µL supernatant was introduced into HPLC for analysis (Emadi-Konjin et al. 2005).