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Specific magnetic isolation for direct detection of HPV16

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Abstract

Finding a suitable DNA purification system is vital for the success of many PCR based diagnostic tests. This report demonstrates the value of magnetic beads in combination with real-time PCR for the sequence-specific isolation and detection of episomal HPV16 DNA. In order to maximize the isolation, two purification procedures were evaluated. Compared to the indirect method, in which the target was magnetically labeled after being hybridized to the capture probes, much higher efficiencies were obtained by directly capturing the target using DNA functionalized beads. These higher efficiencies were obtained by carefully tuning the capture probe density on the beads. When modifying the beads with dual-biotinylated capture probes or introducing beads modified with different capture probes, the amount of HPV16 isolated from spiked clinical swab samples even increased further. This not only resulted in the use of dual-biotinylated capture probes in higher purification efficiencies, but also the thermostability of the DNA-bead linkage was found to improve. In summary, this study shows that DNA functionalized magnetic beads are very promising diagnostic tools as they allow for a specific, simple, and fast isolation and concentration of minute quantities of DNA from complex clinical samples.

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Acknowledgments

The authors would like to thank all members of the FNS research group for their valuable scientific input. Sara Peeters is grateful to the Institute for the Promotion and Innovation through Science and Technology (IWT-Vlaanderen).

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Authors and Affiliations

  1. IMEC, SSET/FNS, Kapeldreef 75, 3001, Leuven, Belgium

    S. Peeters, T. Stakenborg, F. Colle, C. X. Liu & L. Lagae

  2. Department of Microbiology and Immunology, KULeuven, 3001, Leuven, Belgium

    S. Peeters & M. Van Ranst

  3. Department of Biomedical Sciences, University of Antwerp, 2610, Antwerp, Belgium

    F. Colle

  4. Applied Molecular Genomics Group, Department of Molecular Genetics, Flanders Interuniversity Institute for Biotechnology (VIB), Flanders, Belgium

    F. Colle

  5. Department of Physics and Astronomy, KULeuven, 3001, Leuven, Belgium

    L. Lagae

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  1. S. Peeters
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  2. T. Stakenborg
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  3. F. Colle
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  4. C. X. Liu
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  5. L. Lagae
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  6. M. Van Ranst
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Correspondence to S. Peeters.

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Peeters, S., Stakenborg, T., Colle, F. et al. Specific magnetic isolation for direct detection of HPV16. Eur J Clin Microbiol Infect Dis 31, 539–546 (2012). https://doi.org/10.1007/s10096-011-1345-4

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  • DOI: https://doi.org/10.1007/s10096-011-1345-4

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