Specialized for automated, multiplex assays detecting blood-borne viruses for blood bank and organ donor testing.
Low LOD 1-Step RT-qPCR Mix
Specialized for automated, multiplex assays detecting blood-borne viruses for blood bank and organ donor testing
Suitable Application
Blood Banking
Transmission of viral infections through organ transplants and the transfusion of blood products has been known for decades but was considered to be an unavoidable consequence of a life- saving treatment. Molecular testing has helped reduce the number of viral transfusion-transmitted infections due to its superior sensitivity and specificity over other testing methods. However, unique to blood banking and organ donor testing is the challenge of detecting viruses during the window period before seroconversion. PCR inhibitors traditionally found in blood samples have posed a significant challenge in developing molecular assays that can detect low level viremia in early stage infections. Meridian’s new Low LOD 1-Step RT-qPCR Mix has been specifically developed for highly reproducible first-strand cDNA synthesis and qPCR in a single tube. It is formulated with an antibody-mediated hot-start DNA polymerase and uses the latest advances in buffer chemistry and enhancers. Containing dUTP, it is ideal for automated multiplex testing of viruses such as HCV, HBV, HIV-1, HIV-2, HAV and Parvovirus B19. Low LOD 1-Step RT-qPCR Mix has already been successfully adopted for a CE-IVD Marked NAT Blood Banking assay.
Low LOD 1-Step RT-qPCR Mix • Suitable for detecting purified RNA and DNA viruses at very low titers • Novel buffer system optimized for multiplex assays • Perform first-stand DNA synthesis and qPCR in a single tube
PRODUCT
CAT NO.
VOLUME REACTIONS
10 mL
1,000 Rxn
Low LOD 1-Step RT-qPCR Mix
MDX025
100 mL
10,000 Rxn
www.MeridianLifeScience.com
Product Highlights
Figure 1: Sensitive multiplex qPCR for simultaneous detection of DNA and RNA viruses 22 replicates of each extract dilution were tested in multiplex qPCR reactions using the Low LOD 1-Step RT-qPCR Mix and the limit of detection hit rate was determined as a percentage of positive amplifications. The multiplex reactions were HAV, HBV, HCV and an extraction control and HIV-1, HIV-2, Parvovirus B19 and an extraction control. Results illustrate that Low LOD 1-Step RT-qPCR Mix can be used in multiplex reactions with both RNA and DNA viral templates, making it suitable for monitoring viral load for different types of virus. Sensitive and reproducible detection of low-level viremia for both RNA viruses (HCV, HIV-1, HIV-2 and HAV) and DNA viruses (HBV and Parvovirus B19).
HCV (RNA Virus)
HIV-1 (RNA Virus)
HAV (RNA Virus)
100
100
100
75
75
75
50
50
50
25
25
25
0
0
0
150 IU/mL 75 IU/mL 37.5 IU/mL 18.75 IU/mL 9.375 IU/mL 4.68 IU/mL
500 IU/mL 250 IU/mL 125 IU/mL 62.5 IU/mL 31.3 IU/mL 15.6 IU/mL
30 IU/mL
15 IU/mL
7.5 IU/mL 3.75 IU/mL 1.9 IU/mL 0.9 IU/mL
HIV-2 (RNA Virus)
B19 (DNA Virus)
HBV (DNA Virus)
100
100
100
75
75
75
50
50
50
25
25
25
0
0
0
30 IU/mL
15 IU/mL
7.5 IU/mL 3.75 IU/mL 1.9 IU/mL 0.9 IU/mL
500 IU/mL 250 IU/mL 125 IU/mL 62.5 IU/mL 31.3 IU/mL 15.6 IU/mL
60 IU/mL
30 IU/mL
15 IU/mL
7.5 IU/mL 3.75 IU/mL 1.9 IU/mL
Figure 2: Low limit of detection (LOD) for both RNA and DNA virus 24 replicates of purified blood viral extracts were tested in singleplex qPCR reactions using the LowLOD 1-Step RT-qPCR Mix at the indicated IU/mL. The singleplex reactions were HAV, HBV, HCV, HIV-1, HIV-2, Parvovirus B19. The amplification curve results illustrate that Low LOD 1-Step RT-qPCR Mix can be used with both RNA and DNA viruses as low as 5 IU/mL.
Amplification Curves plification Curves HCV (30 IU/mL) Amplification Curves
Amplification Curves HBV (15 IU/mL)
Amplification Curves Amplification Curves HIV-1 (60 IU/mL)
Amplification Curves
23.688 23.079 23.079 23.6 8
63.075
23.688
21.688 21.079 21.079 21.6 8
21.688
57.075
19.688 19.079 19.079 19.6 8
19.688
51.075
17.688 17.079 17.079 17.6 8
17.688
45.075
15.688 15.079 15.079 15.6 8
15.688
39.075
13.688 13.079 13.079 13.6 8
13.688
33.075
11.688 11.079 11.079 1.6 8
11.688
27.075
9.688 9.079 9.079 9.6 8
9.688
21.075
7.688 7.079 7.079 7.6 8
7.688
15.075
5.688 5.079 5.079 3.688 3.079 3.079 3.6 8 1.688 1.079 1.079 1.6 8 5.6 8
5.688
3.688
9.075
1.688
3.075
2 4 6 8 10 2 4 6 8 10 12
12 12
14 14 20 14
16 16
18 18 26
20 20
22 22
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26 26
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30 30
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28
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34
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30
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Amplification Curves Amplification Curves HIV-2 (5 IU/mL)
Amplification Curves HAV (5 IU/mL) Amplification Curves
B19 (60 IU/mL)
Amplification Curves
Amplification Curves
6.628
40.432
43.077
36.932
6.028
39.077
33.432
5.428
35.077
29.932
4.828
31.077
26.432
4.228
27.077
22.932
3.628
23.077
19.432
3.028
19.077
15.932
2.428
15.077
12.432
1.828
11.077
8.932
1.228
7.077
5.432
0.628
3.077
1.932
0.028
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10/19
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