A genotype-independent real-time PCR assay for quantification of hepatitis B virus DNA.
Accurate quantification of hepatitis B virus (HBV) DNA levels is important for monitoring patients with chronic HBV infection and for assessing their responses to antiviral therapy. This study aimed to develop a real-time PCR assay that is sensitive and can accurately quantify a wide range of HBV DNA levels across the known HBV genotypes. An "in-house" real-time PCR assay using primers and a TaqMan probe in a highly conserved region of the HBV surface gene was designed. The assay was standardized against a WHO standard and validated against plasmids of HBV genotypes A through H. The linear quantification range was approximately 5 x 10(0) to 2.0 x 10(9) IU/ml. Results of samples from patients infected with HBV genotypes A through H tested using our real-time "in-house" PCR assay showed an excellent correlation with those of the Cobas Amplicor HBV Monitor (R2=0.9435) and the Cobas TaqMan HBV (R2=0.9873) tests. We have established a real-time PCR assay that is genotype independent and can accurately quantify a wide range of HBV DNA levels. Further studies of additional samples are ongoing to validate the genotype independence of our assay.
Liu
Ying
Y
Division of Gastroenterology, University of Michigan Medical Center, 1500 East Medical Center Dr., 3912 Taubman Center, Box 0362, Ann Arbor, MI 48109-0362, USA.
Hussain
Munira
M
Wong
Stephen
S
Fung
Scott K
SK
Yim
Hyung Joon
HJ
Lok
Anna S F
AS
D017931:DNA Primers; D004279:DNA, Viral / Q000032:analysis* / Q000302:isolation & purification; D005838:Genotype; D006509:Hepatitis B; D006515:Hepatitis B virus / Q000145:classification* / Q000235:genetics*; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D011933:Reagent Kits, Diagnostic; D012680:Sensitivity and Specificity; D019914:Taq Polymerase
D017931:DNA Primers; D004279:DNA, Viral; D011933:Reagent Kits, Diagnostic; D019914:Taq Polymerase