Quantitative PCR for human herpesviruses 6 and 7.
AUTOR(ES)
Secchiero, P
RESUMO
A quantitative PCR assay for the detection of human herpesvirus 6 (HHV-6) (variants A and B) and HHV-7 DNAs in clinical samples was developed. The assay uses a nonhomologous internal standard (IS) for each virus that is coamplified with the wild-type target sequence in the same vial and with the same pair of primers. This method allows for a correction of the variability of efficiency of the PCR technique. A standard curve is constructed for each experiment by coamplification of known quantities of the cloned HHV-6 or HHV-7 target templates with the respective IS. Absolute quantitation of the test samples is then achieved by determining the viral target/IS ratio of the hybridization signals of the amplification products and plotting this value against the standard curve. Using this assay, we quantitated the amount of HHV-6 or HHV-7 DNA in infected cell cultures and demonstrated an inhibitory effect of phosphonoformic acid on the replication of HHV-6 and HHV-7 in vitro. As the first clinical application of this procedure, we performed preliminary measurements of the loads of HHV-6 and HHV-7 in lymph nodes from patients with Hodgkin's disease and AIDS. Application of this quantitative PCR method should be helpful for elucidating the pathogenic roles of HHV-6 and HHV-7.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=228347Documentos Relacionados
- Human herpesviruses 6 and 7 in cervixes of pregnant women.
- In vitro activation of human herpesviruses 6 and 7 from latency.
- Real-Time Quantitative PCR for Human Herpesvirus 6 DNA
- Detection of human herpesviruses 6 and 7 genomic sequences in brain tumours.
- Sensitive Method for Detection of Human Herpesviruses 6 and 7 in Saliva Collected in Field Studies