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Reproducibility of a high-throughput HIV-1 genotypic resistance assay over time (2001–2007)

Background

Designed for innovative HIV-1 diagnostics and resistance analysis, a high-throughput laboratory-developed (Protease-Reverse-Transcriptase) genotyping assay was implemented at Virco (Belgium). Following regulatory guidelines, assay variability was monitored over time by means of a quarterly internal quality control (QC) panel.

Methods

The panel comprises seven well-characterized recombinant viruses that were derived from clinical isolates. Virus stocks were grown and later re-cultured as needed and aliquoted for use at each quarterly testing between Q4 in 2001 through Q1 in 2007. The nucleotide similarity among the original virus stocks and subsequent re-cultured viruses, was determined using pairwise BLAST analysis (BioEdit Sequence Alignment Editor).

Summary of results

A total of 603 and 867 pairwise BLAST comparisons were made, yielding an average within and between original and re-cultured virus stocks similarity of 99.60% ± 0.27 and 99.52% ± 0.33, respectively.

Conclusion

Since both BLAST values did not differ, it can be concluded that re-culturing of viruses did not lead to a significant decrease in similarity. Furthermore, despite the presence of high levels of nucleotide sequence mixtures of the original clinical isolates, the high reproducibility of the genotyping assay (more than 99% sequence identity) was demonstrated. A combination of optimal assay design, rigorous personnel training, the testing of the internal QC panel at regular intervals and participation in a CLIA-approved 6-monthly external proficiency testing program (Accutest) ensures that quality standards are consistently met for the genotyping assay performed at Virco.

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Open Access This article is published under license to BioMed Central Ltd. This is an Open Access article is distributed under the terms of the Creative Commons Attribution 2.0 International License (https://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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Deloof, A., Muyldermans, G., Pattery, T. et al. Reproducibility of a high-throughput HIV-1 genotypic resistance assay over time (2001–2007). JIAS 11, P205 (2008). https://doi.org/10.1186/1758-2652-11-S1-P205

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  • DOI: https://doi.org/10.1186/1758-2652-11-S1-P205

Keywords

  • Proficiency Testing
  • Genotyping Assay
  • Virus Stock
  • Internal Quality Control
  • BioEdit Sequence Alignment Editor