Table 1. Limit of Detection of HCV WHO International Standard
Concentration (IU/ML)
|
Reactive
|
Non-Reactive
|
Invalid
|
Total
|
Percent Valids Detected
|
95% CI*
|
0.1
|
1
|
59
|
0
|
60
|
1.7
|
0.3–8.9
|
1
|
23
|
36
|
1
|
60
|
39.0
|
27.6–51.7
|
2.5
|
178
|
61
|
1
|
240
|
74.5
|
68.7–79.7
|
5
|
220
|
19
|
1
|
240
|
92.1
|
87.9–94.9
|
7.5
|
236
|
2
|
2
|
240
|
99.2
|
97.0–99.8
|
10
|
60
|
0
|
0
|
60
|
100
|
94.0–100
|
18.5
|
59
|
0
|
1
|
60
|
100
|
94.0–100
|
50
|
60
|
0
|
0
|
60
|
100
|
94.0–100
|
The Limit of Detection (LOD) of the Aptima HCV RNA Qualitative Assay was determined by testing serial dilutions of the WHO International Standard for HCV genotype 1 RNA (NIBSC code 96/790).6 Each panel member was tested in replicates ranging from 60 to 240. Serial dilutions of the WHO International Standard for HCV genotype 1 RNA were detected 95% of the time as low as 7.5 IU/mL. Linear regression analysis determined 5.3IU/mL ≥(95% probability) as the limit of detection for the Aptima HCV RNA qualitative assay.
Table 2. HCV Genotype Detection (Transcripts)
HCV Genotype
|
Concentration (IU/ML) (copies/mL)
|
Reactive
|
Non-Reactive
|
Invalid
|
Total
|
Percent Valids Detected
|
95% CI*
|
1
|
9.6 (50)
|
716
|
3
|
1
|
720
|
99.6
|
98.8–99.9
|
2a
|
9.6 (50)
|
357
|
3
|
0
|
360
|
99.2
|
97.6–99.7
|
2b
|
14.4 (75)
|
350
|
10
|
0
|
360
|
97.2
|
95.0–98.5
|
3a
|
9.5 (50)
|
356
|
4
|
0
|
360
|
98.9
|
97.2–99.6
|
4a
|
9.6 (50)
|
357
|
1
|
2
|
360
|
99.7
|
98.4–99.9
|
5a
|
9.6 (50)
|
356
|
4
|
0
|
360
|
98.9
|
97.2–99.6
|
6a
|
9.6 (50)
|
685
|
35
|
0
|
720
|
95.1
|
93.3–96.5
|
* Confidence Interval
Note: Invalid results were not included in the data calculations for Percent Valids Detected.
Transcripts of HCV genotypes 1, 2a, 2b, 3a, 4a, 5a, and 6a made from the 5’-untranslated region of the HCV genome were tested using the Aptima HCV RNA qualitative assay. All transcripts were quantitated using phosphate analysis and confirmed using hyperchromicity and OD260. The copies/mL were converted to IU/mL using an in-house conversion factor 5.2 copies/mL = 1IU/mL. Dilutions of each transcript were tested at 9.6 IU/mL (50 copies/mL) for genotypes 1, 2a, 3a, 4a, 5a, and 6a, and at 14.4 IU/mL (75 copies/mL) for HCV2b. Each transcript was tested in replicates ranging from 360 to 720.
Table 3. HCV Genotype Detection (Clinical Specimens)
HCV Genotype
|
Concentration (IU/ML) (copies/mL)
|
Total
|
Reactive
|
Non-Reactive
|
Invalid
|
Percent Valids Detected
|
95% CI*
|
1
|
4.8 (25)
|
90
|
87
|
3
|
0
|
96.7
|
90.7–98.9
|
1
|
9.6 (50)
|
495
|
490
|
4
|
1
|
99.2
|
97.9–99.7
|
1
|
14.4 (75)
|
60
|
59
|
1
|
0
|
98.3
|
91.1–99.7
|
1
|
19.2 (100)
|
90
|
90
|
0
|
0
|
100
|
95.9–100
|
1
|
96.2 (500)
|
60
|
60
|
0
|
0
|
100
|
94.0–100
|
1
|
9,615.4 (50,000)
|
30
|
30
|
0
|
0
|
100
|
88.7–100
|
2b
|
9.6 (50)
|
210
|
200
|
10
|
0
|
95.2
|
91.5–97.4
|
2b
|
14.4 (75)
|
176
|
173
|
3
|
0
|
98.3
|
95.1–99.4
|
2b
|
19.2 (100)
|
120
|
120
|
0
|
0
|
100
|
96.9–100
|
2b
|
28.8 (150)
|
30
|
29
|
0
|
1
|
100
|
88.3–100
|
2b
|
38.5 (200)
|
90
|
90
|
0
|
0
|
100
|
95.9–100
|
2b
|
57.7 (300)
|
10
|
10
|
0
|
0
|
100
|
72.3–100
|
3a
|
9.6 (50)
|
120
|
120
|
0
|
0
|
100
|
96.7–100
|
3a
|
19.2 (100)
|
60
|
60
|
0
|
0
|
100
|
94.0–100
|
3a
|
38.5 (200)
|
40
|
40
|
0
|
0
|
100
|
91.2–100
|
4/4a
|
9.6 (50)
|
120
|
114
|
6
|
0
|
95.0
|
89.5–97.7
|
4/4a
|
14.4 (75)
|
30
|
30
|
0
|
0
|
100
|
88.7–100
|
5a
|
9.6 (50)
|
120
|
118
|
0
|
2
|
100
|
96.9–100
|
6a
|
9.6 (50)
|
120
|
114
|
3
|
3
|
97.4
|
92.7–99.1
|
6a
|
14.4 (75)
|
30
|
30
|
0
|
0
|
100
|
88.7–100
|
* Confidence Interval
Note: Invalid results were not included in the data calculations for Percent Valids Detected.
Clinical specimens representing HCV genotypes 1 to 6 at different concentrations were used to determine the percent detection of the Aptima HCV RNA qualitative assay. The specimens were quantitated using the VERSANT HCV RNA 3.0 assay (bDNA). The genotypes of the specimens were provided by the specimen vendor and confirmed using the VERSANT HCV Genotype assay (LiPA) and sequencing. The results are shown in Table 3.The overall percent detected across all HCV genotypes tested was ≥95% at 9.6 IU/mL (50 copies/mL).
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