Cross-Platform Evaluation of Commercial Real-Time SYBR Green RT-PCR Kits for Sensitive and Rapid Detection of European Bat Lyssavirus Type 1
Table 5
Analysis of reproducibility of one-step and two-step RT-PCR. (a) shows for each two-step kit, the intra- and interassay reproducibility for six dilutions of EBLV-1 tested eight times in each of three independent runs ( = 144 samples) and (b) shows the inter- and intra-assay reproducibility for all tested one-step kits.
(a)
2-step methods
Kit
Number of copies/sample
Intra-assay variability
Inter-assay variability
Run 1
Run 2
Run 3
Average
Average Ct ± SD Ct
CV (%)
Number of pos.
Average Ct ± SD Ct
CV (%)
Number of pos.
Average Ct ± SD Ct
CV (%)
Number of pos.
Ct ± SD Ct
CV (%)
Average Ct ± SD Ct
CV (%)
IScript RT + qPCR SsoFast
100
30.46 ± 0.41
1.35
8
30.08 ± 0.57
1.90
8
29.82 ± 0.41
1.38
8
30.12 ± 0.46
1.54
30.12 ± 0.52
1.74
50
31.11 ± 0.93
2.99
6
31.84 ± 1.01
3.19
6
30.80 ± 0.46
1.51
8
31.25 ± 0.80
2.56
31.20 ± 0.88
2.83
25
32.49 ± 0.59
1.83
6
31.60 ± 0.67
2.12
8
31.27 ± 0.68
2.19
6
31.79 ± 0.65
2.04
31.77 ± 0.80
2.51
10
32.38 ± 0.91
2.81
6
32.69 ± 1.19
3.63
6
31.97 ± 0.76
2.39
5
32.35 ± 0.95
2.94
32.37 ± 0.97
2.99
5
33.14
—
1
—
—
0
31.98 ± 0.05
0.15
3
32.56 ± 0.05
0.15
32.27 ± 0.58
1.81
0.5
32.17
—
1
—
—
0
32.11 ± 0.76
2.36
3
32.14 ± 0.76
2.36
32.13 ± 0.62
1.93
250
30.71 ± 0.49
1.61
8
30.70 ± 0.39
1.28
8
30.61 ± 0.33
1.08
8
30.67 ± 0.41
1.32
30.67 ± 0.40
1.29
100
32.30 ± 0.74
2.28
8
32.03 ± 0.90
2.81
8
32.19 ± 0.40
1.24
8
32.17 ± 0.68
2.11
32.17 ± 0.69
2.14
50
34.02 ± 0.93
2.73
6
33.24 ± 0.48
1.44
8
33.16 ± 0.93
2.87
8
33.47 ± 0.79
2.35
33.42 ± 0.85
2.55
25
34.63 ± 1.02
2.95
8
33.78 ± 0.75
2.23
6
34.24 ± 0.88
2.58
6
34.21 ± 0.89
2.59
34.25 ± 0.93
2.72
10
35.95 ± 1.12
3.12
5
35.26 ± 0.47
1.34
5
34.68 ± 00.63
1.80
6
35.30 ± 0.74
2.09
35.26 ± 0.90
2.56
5
33.84 ± 0.62
1.84
3
—
—
0
—
—
0
34.83 ± 0.60
1.71
33.84 ± 0.62
1.84
QuantiTect RT + qPCR Quantitect
250
31.37 ± 0.69
2.20
8
31.45 ± 0.52
1.65
8
31.22 ± 0.46
1.47
8
31.35 ± 0.56
1.77
31.35 ± 0.55
1.75
100
32.46 ± 0.80
2.46
7
32.72 ± 0.92
2.81
7
33.90 ± 0.90
2.65
7
33.03 ± 0.87
2.64
33.03 ± 1.05
3.18
50
33.59 ± 0.60
1.79
4
34.34 ± 2.32
6.75
5
33.66 ± 0.79
2.35
6
33.86 ± 1.24
3.63
33.87 ± 1.40
4.13
25
32.98 ± 0.74
2.24
4
33.58 ± 1.18
3.51
5
34.12 ± 2.07
6.08
4
33.56 ± 1.33
3.94
33.56 ± 1.38
4.10
10
33.54 ± 0.18
0.54
3
33.86 ± 0.61
1.80
2
34.73 ± 0.62
1.79
2
34.04 ± 0.47
1.38
33.97 ± 0.65
1.92
5
—
—
0
35.52
—
1
—
—
0
35.52
—
35.52
—
500
34.09 ± 0.36
1.35
8
36.37 ± 1.18
3.26
6
32.97 ± 0.38
1.16
8
34.30 ± 1.54
1.92
34.30 ± 1.54
4.48
250
36.42 ± 0.46
6.43
8
33.66 ± 0.80
2.39
8
33.61 ± 0.83
2.46
8
34.56 ± 1.97
3.76
34.56 ± 1.97
5.69
100
34.39 ± 2.34
2.57
8
34.60 ± 0.98
2.84
8
34.50 ± 0.91
2.65
8
34.50 ± 0.89
2.69
34.50 ± 0.89
2.58
50
36.28 ± 0.88
4.75
6
35.07 ± 0.91
2.59
7
34.92 ± 1.04
2.97
7
35.38 ± 1.32
3.44
35.38 ± 1.32
3.74
25
35.97 ± 1.72
2.30
5
36.28 ± 0.88
2.42
4
36.44 ± 0.78
2.14
5
36.23 ± 0.79
2.29
36.23 ± 0.79
2.17
10
—
—
0
—
—
0
37.65 ± 0.79
2.10
2
37.65 ± 0.79
2.10
37.65 ± 0.79
2.10
Maxima RT + qPCR Maxima
100
31.83 ± 0.84
2.63
31.38 ± 0.22
0.70
30.09 ± 0.41
1.37
31.1 ± 0.49
1.57
31.1 ± 0.92
2.95
50
33.35 ± 1.09
3.26
8
33.42 ± 0.88
2.64
5
31.12 ± 0.64
2.05
8
32.63 ± 0.87
2.65
32.51 ± 1.40
4.32
25
33.67 ± 0.88
2.61
5
32.77 ± 1.12
3.40
6
32.47 ± 0.72
2.21
7
32.97 ± 0.90
2.74
32.90 ± 0.99
3.02
10
37.16 ± 4.64
12.49
3
34.00 ± 1.41
4.16
4
34.35 ± 1.33
3.88
5
35.17 ± 2.46
6.84
34.94 ± 2.63
7.54
5
35.44 ± 0.24
0.68
2
38.65
—
1
34.58 ± 1.24
3.57
3
36.22 ± 0.74
2.12
35.55 ± 1.76
4.96
2.5
—
—
0
33.14
—
1
33.47 ± 1.76
5.26
2
33.30 ± 1.76
5.26
33.36 ± 1.26
3.77
250
29.28 ± 0.58
1.99
8
29.05 ± 0.38
1.30
8
29.04 ± 0.39
1.35
8
29.12 ± 0.45
1.55
29.12 ± 0.45
1.56
100
30.53 ± 0.56
1.85
8
30.17 ± 0.48
1.59
8
30.31 ± 0.43
1.43
8
30.34 ± 0.50
1.62
30.34 ± 0.50
1.64
50
31.54 ± 1.25
3.97
8
31.88 ± 1.37
4.29
8
31.72 ± 0.56
1.77
8
31.71 ± 1.08
3.34
31.71 ± 1.08
3.40
25
33.00 ± 1.15
3.48
6
32.38 ± 0.79
2.43
7
32.38 ± 0.82
2.54
7
32.56 ± 0.92
2.82
32.56 ± 0.92
2.81
10
33.08 ± 0.64
1.93
3
33.02 ± 1.38
4.17
4
33.40 ± 1.46
4.38
4
33.17 ± 1.15
3.49
33.17 ± 1.15
3.47
50
33.03 ± 0.15
0.45
2
33.55
—
1
32.99
—
1
33.15 ± 0.28
0.45
33.15 ± 0.28
0.85
qScript RT + qPCR Perfecta
250
27.04 ± 0.24
0.88
8
26.75 ± 0.26
0.98
8
26.77 ± 0.24
0.90
8
26.85 ± 0.25
0.92
26.85 ± 0.27
1.01
100
30.54 ± 0.54
1.76
8
29.84 ± 0.54
1.82
8
30.46 ± 1.07
3.50
8
30.28 ± 0.72
2.36
30.28 ± 0.79
2.61
50
32.03 ± 2.90
9.05
7
30.72 ± 0.89
2.92
8
30.78 ± 0.82
2.67
8
31.18 ± 1.54
4.88
31.14 ± 1.77
5.68
25
32.50 ± 0.98
3.02
7
32.41 ± 1.33
4.10
7
33.63 ± 2.16
6.42
5
32.85 ± 1.49
4.52
32.76 ± 1.49
4.56
10
32.22 ± 0.81
2.52
5
32.87 ± 0.46
1.39
3
32.70 ± 0.56
1.71
2
32.60 ± 0.61
1.87
32.51 ± 0.69
2.11
5
32.78 ± 0.15
0.45
2
—
—
0
33.56 ± 1.55
4.61
2
33.18 ± 0.85
2.53
33.18 ± 1.00
3.03
250
29.47 ± 0.22
0.74
8
29.49 ± 0.54
1.83
7
29.82 ± 0.38
1.29
8
29.60 ± 0.41
1.29
29.60 ± 0.41
1.39
100
30.99 ± 1.09
3.52
8
31.14 ± 1.26
4.04
8
31.02 ± 0.52
1.69
8
31.05 ± 0.96
3.08
31.05 ± 0.96
3.11
50
31.87 ± 0.60
1.88
8
31.37 ± 0.57
1.80
8
33.13 ± 1.12
3.37
8
32.12 ± 1.08
2.35
32.12 ± 1.08
3.35
25
33.35 ± 2.99
8.96
7
32.49 ± 0.68
2.11
8
32.71 ± 0.87
2.65
8
32.83 ± 1.72
4.57
32.83 ± 1.72
5.24
10
33.00 ± 0.49
1.47
5
33.45 ± 0.18
0.53
2
32.98 ± 0.74
2.25
4
33.07 ± 0.54
1.42
33.07 ± 0.54
1.65
5
33.04 ± 1.01
3.06
2
33.03 ± 0.74
2.23
2
33.59
—
1
33.14 ± 0.67
2.65
33.14 ± 0.67
2.03
SS Vilo RT + qPCR Express
100
27.68 ± 0.59
2.13
8
27.82 ± 0.55
1.97
8
27.59 ± 0.29
1.06
8
27.70 ± 0.48
1.72
27.70 ± 0.48
1.74
50
29.42 ± 0.62
2.11
8
29.27 ± 0.54
1.85
8
29.21 ± 0.60
2.06
8
29.30 ± 0.59
2.01
29.30 ± 0.57
1.95
25
30.38 ± 0.67
2.21
6
30.22 ± 1.24
4.11
8
29.60 ± 0.48
1.62
8
30.07 ± 0.80
2.64
30.04 ± 0.90
3.00
10
31.36 ± 0.49
1.58
7
31.30 ± 0.71
2.27
7
31.92 ± 0.78
2.45
6
31.53 ± 0.66
2.10
31.51 ± 0.69
2.19
5
31.49 ± 0.53
1.68
4
31.63 ± 0.98
3.08
4
31.64
—
1
31.59 ± 0.75
2.38
31.57 ± 0.68
2.16
0.5
—
—
0
—
—
0
31.59
—
1
31.59
—
31.59
—
100
31.78 ± 0.53
1.67
8
30.13 ± 0.32
1.07
8
30.75 ± 0.65
2.11
8
30.89 ± 0.85
1.62
30.89 ± 0.85
2.76
50
32.96 ± 0.42
1.29
8
31.34 ± 0.50
1.61
8
31.49 ± 0.97
3.07
8
31.93 ± 0.99
1.99
31.93 ± 0.99
3.09
25
33.72 ± 1.05
3.10
7
32.15 ± 0.98
3.06
8
32.09 ± 0.22
0.69
6
32.66 ± 1.13
2.28
32.66 ± 1.13
3.46
10
34.44 ± 0.60
1.74
7
33.61 ± 0.46
1.35
5
33.30 ± 0.43
1.29
6
33.83 ± 0.71
1.46
33.83 ± 0.71
2.09
5
35.21 ± 0.53
1.51
7
32.64 ± 0.36
1.11
5
32.99 ± 0.06
0.17
2
33.97 ± 1.35
0.93
33.97 ± 1.35
3.98
2.5
39.56 ± 1.86
4.71
3
34.35
1
33.55 ± 0.57
1.69
3
36.24 ± 3.32
3.20
36.24 ± 3.32
9.16
The intra- and interassay variability and coefficient of variations were calculated by the formulas: inter-assay CV = standard deviation of all replicates/(mean Ct-value of all replicates) × 100; intra-assay CV = mean of the CVs of all runs. Average Ct values: mean ± standard deviation. CV (%) = standard deviation/mean Ct × 100. All PCR comparisons were run on the Rotor Gene Q MDx thermocycler (in italic font) and on the Mx3005P thermocycler (in bold).
(b)
1 step methods
Kit
Number of copies/sample
Intra-assay variability
Inter-assay variability
Run 1
Run 2
Run 3
Average
Average Ct ± SD Ct
CV
Number of pos.
Average Ct ± SD Ct
CV
Number of pos.
Average Ct ± SD Ct
CV
Number of pos.
Ct ± SD Ct
CV
Average Ct ± SD Ct
CV
Quantitect 1 step
50
29.76 ± 0.64
2.16
8
28.94 ± 0.55
1.89
8
29.27 ± 0.23
0.79
8
29.32 ± 0.47
1.61
29.32 ± 0.59
1.06
25
30.90 ± 0.88
2.86
8
29.84 ± 1.09
3.65
8
30.98 ± 0.76
2.45
8
30.57 ± 0.91
2.99
30.57 ± 1.03
2.02
10
31.72 ± 1.04
3.26
7
32.05 ± 1.06
3.32
7
32.30 ± 0.61
1.88
8
32.03 ± 0.90
2.82
32.04 ± 0.90
3.36
5
33.34 ± 1.20
3.60
6
32.38 ± 0.74
2.28
7
32.85 ± 0.84
2.55
7
32.86 ± 0.92
2.81
32.83 ± 0.96
2.81
2.5
32.51 ± 0.60
1.85
4
33.90 ± 0.33
0.96
4
34.45 ± 1.52
4.42
5
33.62 ± 0.82
2.41
33.68 ± 1.27
3.77
1
33.38 ± 0.96
2.88
4
32.03 ± 0.26
0.80
3
32.92 ± 0.47
1.43
5
32.77 ± 0.56
1.70
32.85 ± 0.79
2.42
50
30.92 ± 0.47
1.52
8
31.23 ± 1.29
0.40
8
31.11 ± 1.32
0.41
8
31.08 ± 1.38
0.43
31.08 ± 0.43
1.38
25
31.77 ± 0.73
2.29
8
31.92 ± 1.49
0.48
8
31.91 ± 0.93
0.30
8
31.87 ± 1.57
0.50
31.87 ± 0.51
1.61
10
32.60 ± 0.47
1.44
8
32.81 ± 0.61
1.87
5
32.49 ± 0.65
1.99
8
32.63 ± 0.58
1.77
32.63 ± 0.57
1.75
5
33.69 ± 0.85
2.53
7
33.82 ± 2.91
0.98
5
33.28 ± 2.50
0.83
6
33.60 ± 2.65
0.89
33.57 ± 0.86
2.57
2.5
34.36 ± 0.73
2.14
5
33.65 ± 1.88
0.63
2
33.27 ± 1.88
0.62
3
33.76 ± 1.96
0.66
33.80 ± 0.80
2.36
1
34.55 ± 1.77
5.13
3
34.97 ± 1.17
0.41
5
33.92 ± 3.41
1.16
1
34.48 ± 3.24
1.11
34.45 ± 1.37
3.97
Rotor Gene 1 step
1000
26.40 ± 0.45
1.69
8
23.74 ± 0.40
1.67
8
27.01 ± 1.18
4.38
8
25.72 ± 1.62
2.58
25.72 ± 1.62
6.32
500
27.49 ± 0.46
1.66
8
24.29 ± 0.34
1.39
8
27.42 ± 1.04
3.78
8
26.40 ± 1.66
2.28
26.40 ± 1.66
6.28
250
28.73 ± 0.76
2.65
8
24.90 ± 0.49
1.96
8
29.27 ± 0.60
2.04
7
27.56 ± 2.09
2.22
27.56 ± 2.09
7.58
100
30.70 ± 0.21
0.67
3
25.50
—
1
31.19 ± 0.66
2.11
2
30.00 ± 2.24
1.39
30.00 ± 2.24
7.46
50
30.31
—
1
—
—
0
—
—
0
30.31
—
30.31
—
25
31.12 ± 0.50
1.61
2
—
—
0
—
—
0
32.12 ± 0.50
1.61
32.12 ± 0.50
1.61
50
28.87 ± 0.99
3.41
8
29.25 ± 0.74
2.52
8
27.97 ± 0.65
2.33
8
28.70 ± 0.79
2.76
28.70 ± 0.94
3.29
25
29.49 ± 1.01
3.44
8
30.36 ± 0.70
2.29
8
28.57 ± 0.39
1.36
8
29.47 ± 0.70
2.36
29.47 ± 1.03
3.50
10
30.29 ± 1.20
3.98
8
30.61 ± 1.44
4.70
8
28.76 ± 0.40
1.39
8
29.89 ± 1.01
3.36
29.89 ± 1.34
4.50
5
30.82 ± 1.06
3.43
6
31.47 ± 1.30
4.12
6
29.90 ± 0.40
1.35
8
30.73 ± 0.92
2.97
30.65 ± 1.12
3.66
2.5
30.87 ± 1.14
3.68
6
31.69 ± 0.57
1.81
5
29.10 ± 0.37
1.26
5
30.55 ± 0.69
2.25
30.57 ± 1.31
4.29
1
31.22
—
1
31.53
—
1
32.01 ± 0.44
1.37
5
31.59 ± 0.44
1.37
31.83 ± 0.48
1.51
Verso 1 step
500
26.50 ± 0.18
0.68
8
26.56 ± 0.14
0.53
8
26.79 ± 0.23
0.85
8
26.61 ± 0.18
0.69
26.61 ± 0.22
0.82
250
27.43 ± 0.18
0.65
8
27.96 ± 0.27
0.98
8
27.89 ± 0.36
1.27
8
27.76 ± 0.27
0.97
27.76 ± 0.36
1.30
100
28.94 ± 0.50
1.72
8
29.42 ± 0.53
1.80
8
29.91 ± 1.18
3.96
8
29.43 ± 0.74
2.49
29.43 ± 0.87
2.95
50
30.49 ± 0.44
1.43
7
31.38 ± 0.86
2.74
8
31.29 ± 1.52
4.87
8
31.05 ± 0.94
3.01
31.08 ± 1.09
3.50
25
32.82 ± 1.14
3.46
7
32.90 ± 0.52
1.57
5
32.50 ± 0.92
2.82
7
32.74 ± 0.86
2.62
32.72 ± 0.90
2.74
10
33.42 ± 0.97
2.91
3
35.22 ± 1.05
2.99
2
34.42
—
1
34.35 ± 1.01
2.95
34.19 ± 1.18
3.44
250
29.39 ± 0.48
1.64
8
29.40 ± 0.23
0.79
8
29.10 ± 0.53
1.83
8
29.20 ± 0.42
1.42
29.30 ± 0.44
1.50
100
30.55 ± 0.25
0.80
8
30.32 ± 0.38
1.25
8
30.02 ± 0.24
0.82
6
30.29 ± 0.29
0.96
30.32 ± 0.36
1.19
50
31.16 ± 0.50
1.59
8
31.10 ± 0.52
1.68
8
30.87 ± 0.42
1.35
7
31.04 ± 0.48
1.54
31.05 ± 0.48
1.53
25
31.71 ± 0.32
1.02
7
31.61 ± 0.42
1.34
8
31.35 ± 0.58
1.84
8
31.56 ± 0.44
1.40
31.55 ± 0.46
1.47
10
31.86 ± 0.54
1.69
7
31.76 ± 0.29
0.91
7
31.65 ± 0.64
2.01
4
31.76 ± 0.49
1.54
31.77 ± 0.46
1.45
5
33.09 ± 0.98
2.97
5
33.13
1
33.43 ± 0.74
2.23
4
33.22 ± 0.86
2.60
33.26 ± 0.80
2.39
qScript 1 step
1000
23.24 ± 0.12
0.53
8
22.97 ± 0.24
1.03
8
23.21 ± 0.09
0.37
8
23.14 ± 0.15
0.64
23.14 ± 0.20
0.86
750
23.64 ± 0.21
0.88
8
23.29 ± 0.08
0.35
8
23.52 ± 0.28
1.21
8
23.48 ± 0.19
0.81
23.48 ± 0.25
1.06
500
24.46 ± 0.32
1.32
8
24.06 ± 0.24
0.98
8
24.07 ± 0.29
1.22
8
24.19 ± 0.28
1.17
24.19 ± 0.33
1.37
250
25.38 ± 0.27
1.07
8
24.93 ± 0.26
1.05
8
25.14 ± 0.34
1.37
8
25.15 ± 0.29
1.16
25.15 ± 0.34
1.35
100
26.26 ± 0.31
1.16
8
25.79 ± 0.24
0.93
7
25.95 ± 0.47
1.80
5
26.00 ± 0.34
1.30
26.02 ± 0.38
1.46
50
—
—
0
—
—
0
26.45 ± 0.18
0.70
2
26.45 ± 0.18
0.70
26.45 ± 0.18
0.70
2500
28.13 ± 0.83
2.94
8
27.78 ± 0.45
1.60
8
26.32 ± 0.30
1.13
8
27.41 ± 0.52
1.89
27.41 ± 0.97
3.53
1000
28.78 ± 0.09
0.33
8
29.40 ± 1.58
5.39
8
24.89 ± 0.30
1.22
8
27.69 ± 0.66
2.31
27.69 ± 2.23
8.05
750
29.08 ± 0.24
0.82
8
28.92 ± 0.25
0.85
8
28.75 ± 0.11
0.38
8
28.92 ± 0.20
0.68
28.92 ± 0.24
0.84
500
30.69 ± 0.48
1.57
7
29.76 ± 0.55
1.86
8
27.63 ± 0.56
2.01
5
29.36 ± 0.53
1.81
29.66 ± 1.26
4.24
250
31.22 ± 0.23
0.73
7
30.05 ± 0.37
1.24
8
—
—
0
30.64 ± 0.30
0.99
30.60 ± 0.67
2.20
100
—
—
0
30.00
—
1
—
—
0
30.00
—
30.00
—
SSIII Platinium 1 step
1000
24.79 ± 0.24
0.97
8
23.81 ± 0.21
0.89
8
24.46 ± 0.15
0.60
8
24.35 ± 0.20
0.82
24.35 ± 0.46
1.88
750
25.18 ± 0.24
0.95
8
24.30 ± 0.12
0.51
8
24.97 ± 0.10
0.42
8
24.82 ± 0.16
0.63
24.82 ± 0.41
1.67
500
26.66 ± 0.82
3.07
8
25.36 ± 0.23
0.92
8
26.19 ± 0.24
0.92
8
26.07 ± 0.43
1.64
26.07 ± 0.73
2.82
250
28.66 ± 0.86
3.01
8
28.38 ± 1.91
6.72
8
28.97 ± 0.59
2.05
8
28.67 ± 1.12
3.93
28.67 ± 1.22
4.27
100
29.17 ± 0.53
1.80
8
27.78 ± 0.49
1.75
8
28.16 ± 0.53
1.89
7
28.37 ± 0.52
1.82
28.38 ± 0.78
2.76
50
30.95 ± 0.43
1.39
2
29.11 ± 0.67
2.31
4
29.93 ± 1.11
3.70
3
30.00 ± 0.74
2.47
29.79 ± 1.03
3.47
250
26.27 ± 0.26
0.99
8
26.70 ± 0.27
1.02
8
26.67 ± 0.42
1.56
8
26.55 ± 0.37
1.19
26.55 ± 0.37
1.39
100
27.22 ± 0.68
2.48
8
28.02 ± 0.39
1.38
8
27.74 ± 0.34
1.21
8
27.66 ± 0.58
1.69
27.66 ± 0.58
2.09
50
27.56 ± 0.53
1.94
8
28.26 ± 0.41
1.47
8
28.33 ± 0.40
1.41
8
28.05 ± 0.56
1.61
28.05 ± 0.56
2.00
25
27.87 ± 0.69
2.47
8
29.34 ± 0.55
1.89
8
28.58 ± 0.41
1.45
8
28.59 ± 0.82
1.94
28.59 ± 0.82
2.85
10
28.75 ± 0.78
2.70
6
29.44 ± 0.55
1.86
5
29.08 ± 0.33
1.14
7
29.07 ± 0.60
1.90
29.07 ± 0.60
2.07
5
28.82 ± 0.15
0.53
4
28.71 ± 0.21
0.72
3
28.88 ± 0.32
1.12
4
28.81 ± 0.23
0.79
28.81 ± 0.23
0.79
The intra- and interassay variability and coefficient of variations were calculated by the formulas: interassay CV = standard deviation of all replicates/(mean Ct-value of all replicates) × 100, intra-assay CV = mean of the CVs of all runs. Average Ct values: mean ± standard deviation. CV (%) = standard deviation/mean Ct × 100. All PCR comparisons were run on the Rotor Gene Q MDx thermocycler (in italic font) and on the Mx3005P thermocycler (in bold).
