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. Author manuscript; available in PMC: 2023 Jul 24.
Published in final edited form as: Nat Protoc. 2023 Feb 8;18(4):1296–1315. doi: 10.1038/s41596-023-00803-0

Table 5.

Results of the calibration line and the analytical characteristics of the plasma analysis for the EISA-TOF and EISA-Q methods.

EISA-TOF EISA-Q
Internal standard calibration method (0.5–100 μmol/L)a
L-2-HG D-2-HG L-2-HG D-2-HG
Slope ± SD 0.131 ± 0.002 0.116 ± 0.002 0.124 ± 0.002 0.119 ± 0.003
Intercept ± SD 0.011 ± 0.003 0.021 ± 0.003 0.009 ± 0.003 0.013 ± 0.004
R2 0.998 0.997 0.998 0.995
LOQb 0.5 μmol/L 0.5 μmol/L 0.5 μmol/L 0.5 μmol/L
F-value of ANOVAc 3268 2918 4046 1668
Plasma
Trueness % Mean recovery ± SD
10 μmol/L 98 ± 4 100 ± 4 85 ± 6 96 ± 21
50 μmol/L 97 ± 5 98.9 ± 0.1 99 ± 3 104 ± 4
70 μmol/L 95 ± 8 99 ± 10 101 ± 4 104 ± 11
Precision RSD (%)
Between injection repeatability (n=3)d 3.6 2.8 2.3 6.8
Measurement repeatability (n=3)e 2.6 5.5 5.4 5.6
Intermediate Measurement precision (n=9)f 4.1 6.0 4.7 6.7
Recovery (before and after the extraction) % Mean recovery ± SD
106 ± 5 97 ± 4 86 ± 4 88 ± 14
a

Nine standard solutions at concentration levels from 0.5 to 100 μmol/L for L-2-HG and D-2-HG. Each solution contains 10 μmol/L of L-2-HG-d3 and D-2-HG-d3. The calibration line was established by plotting the area ratio of each enantiomer versus the concentration using a 1/x2 weighting.

b

LOQ estimated as the lowest concentration in the calibration that can be quantified with acceptable trueness (between 70–130%) and precision (coefficient of variation <30%).

c

F-value for ANOVA test that confirms that experimental data fit to a linear model. In all cases the p-values were lower than 0.05.

d

Three consecutive injections of a plasma sample spiked with 10 μmol/L of L-2-HG, 10 μmol/L of D-2-HG, 10 μmol/L of L-2-HG-d3 and 10 μmol/L of D-2-HG-d3.

e

Three replicates of plasma samples spiked with 10 μmol/L of L-2-HG, 10 μmol/L of D-2-HG, 10 μmol/L of L-2-HG-d3 and 10 μmol/L of D-2-HG-d3 injected in triplicate on the same day.

f

Three replicates of plasma samples spiked with 10 μmol/L of L-2-HG, 10 μmol/L of D-2-HG, 10 μmol/L of L-2-HG-d3 and 10 μmol/L of D-2-HG-d3 injected during three consecutive days.