Comparative study of conventional and novel strategies for the detection of hepatitis C virus RNA in serum: amplicor, branched-DNA, NASBA and in-house PCR.
The aim of this study was to compare the sensitivity and specificity of conventional procedures (in-house one-stage polymerase chain reaction (PCR) and in-house nested PCR) and of new technologies (rTth DNA polymerase (Amplicor), branched-DNA, NASBA (nucleic acid amplification system)) for the qualitative detection of hepatitis C virus (HCV) RNA in serum of HCV-infected individuals. Serum samples from 37 anti-HCV-positive individuals (15 with a normal alanine aminotransferase (ALT) level, 22 with an elevated ALT level) and 10 anti-HCV-negative individuals as negative controls were studied. A second panel, including 9 diluted serum samples (from 1/10 to 1/100,000) was constituted to establish the differences of sensitivity of the 5 procedures with small quantities of HCV RNA in the serum. The anti-HCV-positive individuals with elevated ALT gave positive results with all 5 procedures. In patients with a normal ALT level, the assays with the highest sensitivity were Amplicor, NASBA and nested RT-PCR, followed by one-stage RT-PCR, then branched-DNA. One false-positive result was observed with Amplicor, and two with in-house nested PCR. On diluted samples, Amplicor, NASBA and nested PCR appeared more sensitive than one-stage PCR and branched-DNA. It is concluded that new procedures have satisfactory sensitivity and specificity and could advantageously replace the conventional PCR procedures for the routine qualitative detection of serum HCV RNA.
Lunel
F
F
Hôpital Pitié-Salpêtrière, Paris, France.
Mariotti
M
M
Cresta
P
P
De la Croix
I
I
Huraux
J M
JM
Lefrère
J J
JJ
D001483:Base Sequence; D017931:DNA Primers; D004279:DNA, Viral; D016174:Hepacivirus / Q000235:genetics / Q000302:isolation & purification*; D006526:Hepatitis C / Q000097:blood / Q000175:diagnosis / Q000821:virology*; D006801:Humans; D008969:Molecular Sequence Data; D021141:Nucleic Acid Amplification Techniques; D016133:Polymerase Chain Reaction / Q000379:methods*; D012367:RNA, Viral / Q000097:blood*; D011933:Reagent Kits, Diagnostic; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D017931:DNA Primers; D004279:DNA, Viral; D012367:RNA, Viral; D011933:Reagent Kits, Diagnostic