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. 1999 Sep;37(9):2793-7.
doi: 10.1128/JCM.37.9.2793-2797.1999.

Automated quantitative analysis of hepatitis B virus DNA by using the Cobas Amplicor HBV monitor test

U Noborg  1 , A Gusdal, E K Pisa, A Hedrum, M Lindh
Affiliations

Automated quantitative analysis of hepatitis B virus DNA by using the Cobas Amplicor HBV monitor test

U Noborg et al. J Clin Microbiol. 1999 Sep.

Abstract

A highly sensitive method of quantitative analysis of hepatitis B virus (HBV) DNA in serum, the Cobas Amplicor HBV Monitor (Cobas-AM) test, was evaluated. Following a manual extraction of viral DNA, amplification, colorimetric detection, and quantitative determination are all automatically performed in the Cobas analyzer. Serially diluted samples with known HBV DNA concentrations were analyzed blindly. All samples with a virus concentration of 400 copies/ml and 83% of samples with a virus concentration of 100 copies/ml could be detected. A linear correlation between input HBV DNA and measured HBV DNA was seen in the range from 100 to 10(5) copies/ml. The mean coefficient of variation was 29.6% for all input levels and 18.9% for HBV DNA concentrations above 400 copies/ml. Samples with an HBV DNA level above 10(9) copies/ml could be reproducibly measured after predilution to 10(-4) or 10(-6) in negative serum; however, the level was underestimated if target DNA after dilution was still above the linear range of the assay. Quantitative results of the Cobas-AM test were interchangeable with measurements by the manual microwell plate version of Amplicor HBV Monitor (MWP-AM); the mean ratio for log Cobas-AM results/log MWP-AM results was 0.97 (standard error of the mean, 0.007) when serum samples from 153 chronic carriers were analyzed. The test should be of value for clinical assessment of chronic carriers and for monitoring the response to antiviral treatment. A limitation is the relatively narrow linear range of the assay, requiring predilution of high-titer (mainly hepatitis B e-antigen-positive) samples.

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Figures

FIG. 1
FIG. 1
Reproducibility and precision of Cobas-AM for detection of HBV DNA at levels in the range from 200 to 100,000 copies/ml. Each sample was analyzed 20 times by each of two technicians; mean values and SEMs are shown.
FIG. 2
FIG. 2
Measurements by Cobas-AM of four high-titer serum samples diluted in negative serum to 1:104 (Dil −4) and 1:106 (Dil −6) prior to analysis. Each sample was analyzed 15 times; mean values and SEMs are shown.
FIG. 3
FIG. 3
Linearity in the detection of HBV DNA in the range from 100 to 200,000 copies/ml by Cobas-AM. Each sample was analyzed six times; mean values and SEMs are shown.
FIG. 4
FIG. 4
Correlation between Cobas-AM and the MTW-AM in the analysis of HBV DNA in serum samples from 153 chronic HBV carriers (36 HBeAg-positive carriers and 118 HBeAg-negative carriers). The mean of repeated (two or three analyses) MWP-AM analyses is plotted against a single value from analysis by Cobas-AM after predilution in negative serum as described in the Materials and Methods section (r = 0.97, b = 1.17).
FIG. 5
FIG. 5
ALT and HBV DNA levels measured by Cobas-AM in a patient with chronic active hepatitis treated with alpha interferon and lamivudine.
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