Quantitative multiplex assay for simultaneous detection of the Indiana serotype of vesicular stomatitis virus and HIV gag.
J Virol Methods
; 143(1): 55-64, 2007 Jul.
Article
en En
| MEDLINE
| ID: mdl-17382412
Assessment of in vivo viral replication of live attenuated recombinant vesicular stomatitis virus (rVSV) vaccine vector candidates encoding HIV gag requires comprehensive preclinical safety studies, and development of sensitive assays to monitor the outcome of vaccination of animals is important. In this study, two 2-step quantitative real-time RT-PCR assays were developed; a singleplex assay to detect VSV genomic RNA from ferrets inoculated intra-cranially (IC) or intra-nasally (IN) with either a wild-type (wt) virus or an attenuated rVSV vector engineered to express HIV gag protein, and a duplex assay to simultaneously detect VSV-N and HIV-gag mRNAs from cynomolgus macaques inoculated intra-thalamically (IT) with the same viruses. Using synthetic oligonucleotides as standards, the lower limit of detection of VSV-N and HIV-gag was 50 copies. Results showed high levels of wt VSV(IN) genomic RNA and mRNA in ferret and macaque tissues, respectively, and significantly lower levels of VSV genomic RNA and VSV-N and HIV-gag mRNAs in tissues from animals inoculated with the attenuated rVSV vector. These assays correlated with both the course of infection for these animals, and the infectious viral load measured by a standard plaque assay, and could be used to determine the safety profile of rVSV vaccine vectors.
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Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
ARN Viral
/
Productos del Gen gag
/
VIH
/
Vacunas contra el SIDA
/
Virus de la Estomatitis Vesicular Indiana
Tipo de estudio:
Diagnostic_studies
Límite:
Animals
Idioma:
En
Revista:
J Virol Methods
Año:
2007
Tipo del documento:
Article
País de afiliación:
Estados Unidos
Pais de publicación:
Países Bajos