Quantification of the relative levels of wild-type and lamivudine-resistant mutant virus in serum of HBV-infected patients using microarray.
During the course of lamivudine administration in hepatitis B virus (HBV)-infected patients, a dynamic development of the viral population in serum is observed. Total HBV level is initially reduced, then lamivudine-resistant mutants appear, and finally, the viral level is increased. All methods of mutant detection so far described can only identify mutants in the serum, and cannot determine the proportion of those mutants. In this paper, we report the development of a novel technique that can quantify the relative proportion of mutants in serum utilizing gene microarray technology. Based on the nucleotide sequence at the loci of the mutations in lamivudine-resistant HBV mutants, 28 probes were designed and dotted on glass film to prepare the oligonucleotide microarray. Ten standard curves were established by employing 15 reference plasmids as templates. Ten standard math functions were simulated, which allowed quantification of the proportion of mutants in the sample by measuring the value of fluorescent intensity on the microarray. By utilizing the standard math function, the relative proportion of two different mutation sequences in the mixed template could be detected with an error <10%. The HBV-lamivudine oligonucleotide microarray is reliable to quantify the relative proportion of wild-type HBV vs HBV mutants in patient's sera.
Li
Z G
ZG
Beijing Institute of Radiation Medicine, Beijing, China.
Chen
L Y
LY
Huang
J
J
Qiao
P
P
Qiu
J M
JM
Wang
S Q
SQ
D001483:Base Sequence; D015331:Cohort Studies; D004279:DNA, Viral / Q000032:analysis; D024882:Drug Resistance, Viral; D005260:Female; D006509:Hepatitis B / Q000097:blood / Q000188:drug therapy* / Q000235:genetics; D006515:Hepatitis B virus / Q000187:drug effects* / Q000235:genetics*; D006801:Humans; D019259:Lamivudine / Q000627:therapeutic use*; D008297:Male; D008969:Molecular Sequence Data; D009154:Mutation; D020411:Oligonucleotide Array Sequence Analysis; D016133:Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D016896:Treatment Outcome; D019562:Viral Load
D004279:DNA, Viral; D019259:Lamivudine