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Table 2 Multilevel validation of multiplex detox assays by comparing corresponding expression analysis results to published data and the “gold-standard” singleplex RT-qPCR method. n =3 replicates were used for the multiplex detox assays and singleplex RT-qPCR analysis and P-values were calculated by the pair wise fixed reallocation randomization test. For the correlation analysis P-values were calculated by Pearson’s correlation coefficient (r) test

From: Rapid multiplex gene expression assays for monitoring metabolic resistance in the major malaria vector Anopheles gambiae

Gene Samples compared (R vs S) Method Comparison between multiplex and singleplex RT-qPCR assays
Published data Multiplex detox assays Singleplex RT-qPCR
Fold change (P-value) [Reference] Fold change (95% CI, P-value) Fold change (95% CI, P-value) Mean % difference of fold changes (95% CI) r (P-value)
CYP6P3 VK7 vs NG 3.30 (P = 0.013) [42] 3.53 (1.44–8.14) P < 0.001 6.58 (1.68–26.39) P < 0.001 -13.60 (-127–100) 0.703 (P = 0.001)
TIA vs NG 29.5 (P = 0.005) [27] 20.7 (9.30–34.2) P = 0.033 15.1 (6.83–40.9) P < 0.001
AKR vs KIS – [23, 35] 13.6 (5.77–29.4) P = 0.024 15.3 (6.59–32.4) P < 0.001
CYP6M2 VK7 vs NG 1.80 (P = 0.0044) [42] 6.58 (3.92–8.94) P = 0.031 3.73 (1.98–9.48) P < 0.001 2.32 (-116–120) 0.941 (P < 0.001)
TIA vs NG 6.41 (P = 0.015) [27] 4.46 (0.942–21.1) P = 0.059 4.99 (0.72–34.6) P = 0.103
AKR vs KIS – [23, 35] 21.5 (5.36–59.36) P = 0.006 31.3 (13.1–99.0) P < 0.001
CYP9K1 VK7 vs NG 1.40 (P = 0.059) [42] 1.28 (0.958–1.71) P = 0.094 2.94 (0.942–9.20) P = 0.063 -56.47 (-151–38.4) 0.809 (P = 0.003)
TIA vs NG 4.33 (P = 0.007) [27] 2.01 (1.72–2.32) P < 0.001 4.60 (1.80–18.3) P < 0.001
AKR vs KIS – [23, 35] 8.20 (6.78–10.9) P = 0.006 9.28 (2.48–20.9) P = 0.026
CYP6P4 VK7 vs NG 2.11 (P = 0.11) [42] 6.77 (5.20–8.49) P = 0.014 8.97 (7.50–10.7) P < 0.001 39.65 (-123–202) 0.856 (P < 0.001)
TIA vs NG 14.4 (P = 0.0047) [27] 13.0 (7.92–18.4) P < 0.001 8.33 (4.05–17.6) P = 0.033
AKR vs KIS – [23, 35] 8.54 (5.25–12.1) P < 0.001 2.73 (1.23–6.73) P = 0.043
CYP6Z1 RSP vs KIS 3.50 (P < 0.05) [43]; 1.31 (P < 0.05) [44] 3.03 (2.58–3.38) P = 0.036 2.69 (1.80–4.79) P < 0.001 -35.86 (-101–29.3) 0.512 (P = 0.03)
ZANU vs KIS > 2.0 (P < 0.05) [44] 3.27 (1.20–5.23) P = 0.045 3.25 (0.81–12.98) P = 0.095
TIA vs NG 1.21 (P > 0.05) [27] 2.14 (0.625–4.68) P = 0.173 2.52 (0.819–7.76) P = 0.11
VK7 vs NG 1.40 (P = 0.013) [42] 0.874 (0.747–1.02) P = 0.09 3.31 (1.02–10.75) P = 0.050
AKR vs KIS – [23, 35] 6.53 (2.52–9.97) P = 0.041 12.0 (3.55–77.4) P = 0.001
GSTE2 ZANU vs KIS 7.80 (P < 0.05) [45] 3.90 (P < 0.05) [44] 57.8 (52.3–67.5) P = 0.041 22.7 (4.24–328.6) P < 0.001 20.58 (-50.5–91.7) 0.678 (P = 0.003)
RSP vs KIS 2.36 (P < 0.05) [44] 30.7 (26.6–34.3) P = 0.017 16.5 (1.30–161.3) P < 0.001
TIA vs NG 0.649 (P > 0.05) [27] 0.652 (0.406–1.05) P = 0.08 0.735 (0.340–1.58) P = 0.382
VK7 vs NG 0.909 (P = 0.75) [42] 1.30 (0.891–1.90) P = 0.174 1.015 (0.403–2.78) P = 0.899
AKR vs KIS – [23, 35] 27.5 (6.90–34.5) P = 0.034 49.5 (12.5–347.3) P < 0.001
CYP6P1 VK7 vs NG 2.20 (P = 0.048) [42]; 1.70 (P < 0.05) [46] 1.55 (1.21–1.84) P = 0.024 2.64 (1.75–3.33) P < 0.001 -59.84 (-111– -8.5) 0.703 (P = 0.002)
TIA vs NG 1.84 (P = 0.0037) [27] 2.85 (2.56–3.28) P = 0.023 4.48 (2.54–6.91) P < 0.001
AKR vs KIS – [23, 35] 4.24 (0.886–9.79) P = 0.066 10.3 (2.17–44.9) P = 0.036
CYP4G16 VK7 vs NG 1.90 (P < 0.05) [46]; 1.10 (P = 0.59) [42] 2.03 (1.67–2.51) P = 0.019 2.63 (1.75–3.33) P < 0.001 -51.21 (-109–7.1) 0.708 (P < 0.001)
TIA vs NG 0.455 (P > 0.05) [27] 1.20 (0.760–1.75) P = 0.319 2.55 (0.980–10.9) P = 0.052
AKR vs KIS – [23, 35] 2.18 (1.29–3.46) P = 0.023 3.87 (1.03–23.2) P = 0.050
  1. Abbreviations: NG, Ngusso strain; TIA, Tiassalé strain; AKR, Akron strain; KIS, Kisumu strain