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PX-28 A comparison study of the biomerieux nucelisens easyMAG and the QIAGEN QIAsymphony extractors
Posters, 12th ESCV, Istanbul / Journal of Clinical Virology 46 Suppl. 1 (2009) S15–S61 Conclusion: The Access CMV IgG assay provides good agreement with the AxSYM CMV IgG assay. The Access CMV IgG assay displays excellent results as confirmed with the VIDAS CMV IgG assay. The Access CMV IgG assay can be used on the Access systems, the high-throughput UniCel® DxI 800 Immunoassay System and the integrated UniCel DxCi systems. PX-25 Dynabeads MyOne SILANE in molecular diagnostic applications H. Lindstrom *, M. Bosnes, A. Keiserud, D. Ellis. Life Technologies, Invitrogen Dynal, Oslo, Norway Today, novel virus discovery and viral diagnostics are increasingly geared toward molecular diagnostics. Methods enabling efficient screening of large populations on automated platforms are vital for fast discovery and containment of emerging viral threats. The successful development of an automated protocol requires a solid phase for nucleic acid capture which is both functional and yields reproducible results. With this in mind we have developed Dynabeads® MyOne™ SILANE. These highly uniform 1 mm magnetic beads present a silica-like surface that has been optimized for the capture of nucleic acids. Our studies show that the beads offer highly reproducible, very wide and linear dynamic range of nucleic acid capture (from tens of micrograms of genomic DNA to just a few copies of viral genome). Furthermore, an increased magnetic mobility provides improved separation efficiency in viscous samples. Here, we present data on highly efficient viral nucleic acid isolation from serum and whole blood using Dynabeads® MyOne SILANE. The beads also show compatibility with downstream enzymatic detection methods such as qPCR and qRT-PCR, eliminating the necessity to elute the isolated nucleic acid off the solid phase prior to detection. This way, the entire nucleic acid isolate from one sample may be run in a single qPCR reaction, further increasing detection sensitivity and simplifying workflow. Together, these results show that Dynabeads® MyOne SILANE are well suited for automated high-throughput platforms without compromising detection sensitivity and reproducibility. PX-26 Evaluation of the Qiagen QIAsymphony® SP automated extraction system on a range of samples for detection of cytomegalovirus, herpes virus I, II and varacella zoster virus, respiratory viruses and norovirus genotypes I and II A.V. Lee1 *, D.A. Clark2 . 1 Department of Virology, HPA Collaborating Laboratory, Barts and the London NHS Trust, London, UK, 2 Department of Virology, Barts and the London NHS Trust, London, UK The QIAGEN QIAsymphony® SP automated extraction system was compared against the bioM´erieux NucliSens easyMAG using a range of samples types and the nucleic acid extracts tested on “in-house” PCR assays. Four real time PCR tests were used: quantitative assay for cytomegalovirus (CMV); respiratory multiplex (10 virus targets); herpes simplex virus (HSV) 1, HSV 2 and varicella zoster virus (VZV); norovirus genotypes I and II (Noro GI/II). Overall, the QIAsymphony® SP gave similar results to the EasyMAG for each assay. For CMV viral loads, 58/62 samples had a mean difference of −0.051 log10 copies/ml (−1.7–1.21). For the respiratory multiplex, 67/70 samples showed a mean difference of 0.068 cycles (−9.21–11.51). For HSV1/HSV2/VZV, 44/45 samples showed a mean difference of 1.06 cycles (−3.76–4.43). Lastly, for Noro GI/II in 15/ 15 samples, there was a mean difference of 0.25 cycles (−3.29–2.38). In total, 217/ 225 (96.4%) samples tested gave concordant results between the QIAsymphony® SP and EasyMAG extractors. Inter- and intra-run variation was determined for a dilution series of CMV and found to be less than 0.63 log10 copies/ml for 3.20 to 5.20 log10 copies/ml. The QIAsymphony® SP offers enhanced automation over the EasyMAG, within a closed system, potentially decreasing the risk of contamination. The platform also includes a continuous loading function, and the operation requirements are relatively straightforward. Our evaluation of the QIAsymphony® SP showed extraction performance comparable to the easyMAG and based on these findings, we have now introduced the former into our routine diagnostic service.
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PX-27 Performance evaluation of PCR and microarray-based assay for rapid herpesvirus diagnostics K. Moilanen1 , M. H¨akkinen1 , G. Cannon2 , J. Kauppinen3 , R. Lienhard4 , M.-L. Tritten4 , L. Mannonen5 , M. Lappalainen5 , R. Vainionp¨aa¨ 6 , A.-K. J¨arvinen1 *. 1 Mobidiag Ltd, Helsinki, Finland, 2 National Virus Reference Laboratory, University College Dublin, Dublin, Ireland, 3 Eastern Finland Laboratory Centre Joint Authority Enterprise (ISLAB), Kuopio, Finland, 4 Admed Microbiologie, La Chaux-de-Fonds, Switzerland, 5 Department of Virology, Laboratory Services (HUSLAB), Helsinki University Hospital, Helsinki, Finland, 6 Department of Virology, University of Turku, Turku, Finland We conducted a performance evaluation study for the Prove-it™ Herpes assay (Mobidiag) to compare results to those of current PCR-based herpes diagnostics. Prove-it™ Herpes identifies seven herpesviruses (HSV-1, HSV-2, VZV, EBV, CMV, HHV-6, HHV-7) in a single assay in three hours, whereas most current PCR-based commercial or homebrew methods search for only one or few viruses at a time. Altogether, 525 cerebrospinal fluid samples were analyzed by both methods in five laboratories in Finland, Ireland, and Switzerland. For Prove-it Herpes, extracted DNA was amplified by PCR followed by hybridization and detection by Prove-it™ Advisor software according to manufacturer’s instructions. Reference PCRs were performed as determined by laboratories. Discordant results were studied by additional PCRs and DNA sequencing. Prove-it™ Herpes revealed multiple additional viral targets, either alone or in combinations, when compared to reference PCR workflows. In 13 confirmed multi-infections, combinations such as HSV-2 with HHV-7 were detected. HHV-7 was observed in nine cases together with other herpesviruses and in eight cases alone. Prove-it™ Herpes revealed also individual targets not suspected initially by patient’s profile such as HHV-6, EBV and CMV. The final sensitivity and specificity for Prove-it™ Herpes assay were 90% and 98%. These results, especially HHV-7 cases, yield also interesting data for further investigations. Prove-it™ Herpes was considered as a rapid and robust diagnostic platform that was easily implemented into laboratory workflow. The broad target coverage and small sample volume required by the assay could benefit diagnostics, and thus, the treatment of life-threatening infections of the central nervous system.
PX-28 A comparison study of the biomerieux nucelisens easyMAG and the QIAGEN QIAsymphony extractors S.J. Shepherd1 , R.N. Gunson1 , T. Hanselle2 , J. Buckels3 , W.F. Carman1 *. 1 West of Scotland Specialist Virology Centre, Gartnavel General Hospital, Glasgow, Scotland, 2 QIAGEN GmbH, Hilden, Germany, 3 QIAGEN Ltd, Crawley, UK The West of Scotland Specialist Virology Centre (WOSSVC) is equipped with a variety of automated nucleic acid extraction machines including the BioMerieux easyMAG and the recently marketed QIAsymphony. Both machines have been described by their manufacturers as compact and easy to use. A direct comparison between the easyMAG and the QIAsymphony was undertaken to determine the suitability of the QIAsymphony to work in conjunction with and/or replace the easyMAG. A total of 300 (149 postitive/151 negative) respiratory, blood, tissue and swab samples were extracted simultaneously on both the easyMAG and the QIAsymphony. The eluted samples from both extractors were then tested in duplicate by qualitative real-time PCR and the cycle threshold (Ct) values for each extract compared. Overall the QIAsymphony performed similarly to the easyMAG, irrespective of sample type or pathogen listed. A small number of differences were seen between some samples. In most cases this was likely to be was related to the low level of virus present within the sample material. The main advantages of the QIAsymphony over the easyMAG are not only the benefit of extracting a larger number of samples per run but also the automated addition and elution of samples, the easyMAG requires both these steps to be carried out manually. The main disadvantage found with the QIAsymphony was the requirement for a specific set volume of starting sample, where the easyMAG can use volumes <200ml, the virus/bacteria kit of the QIAsymphony requires a volume in excess of 200ml.
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PX-27 Performance evaluation of PCR and microarray-based assay for rapid herpesvirus diagnostics K. Moilanen1 , M. H¨akkinen1 , G. Cannon2 , J. Kauppinen3 , R. Lienhard4 , M.-L. Tritten4 , L. Mannonen5 , M. Lappalainen5 , R. Vainionp¨aa¨ 6 , A.-K. J¨arvinen1 *. 1 Mobidiag Ltd, Helsinki, Finland, 2 National Virus Reference Laboratory, University College Dublin, Dublin, Ireland, 3 Eastern Finland Laboratory Centre Joint Authority Enterprise (ISLAB), Kuopio, Finland, 4 Admed Microbiologie, La Chaux-de-Fonds, Switzerland, 5 Department of Virology, Laboratory Services (HUSLAB), Helsinki University Hospital, Helsinki, Finland, 6 Department of Virology, University of Turku, Turku, Finland We conducted a performance evaluation study for the Prove-it™ Herpes assay (Mobidiag) to compare results to those of current PCR-based herpes diagnostics. Prove-it™ Herpes identifies seven herpesviruses (HSV-1, HSV-2, VZV, EBV, CMV, HHV-6, HHV-7) in a single assay in three hours, whereas most current PCR-based commercial or homebrew methods search for only one or few viruses at a time. Altogether, 525 cerebrospinal fluid samples were analyzed by both methods in five laboratories in Finland, Ireland, and Switzerland. For Prove-it Herpes, extracted DNA was amplified by PCR followed by hybridization and detection by Prove-it™ Advisor software according to manufacturer’s instructions. Reference PCRs were performed as determined by laboratories. Discordant results were studied by additional PCRs and DNA sequencing. Prove-it™ Herpes revealed multiple additional viral targets, either alone or in combinations, when compared to reference PCR workflows. In 13 confirmed multi-infections, combinations such as HSV-2 with HHV-7 were detected. HHV-7 was observed in nine cases together with other herpesviruses and in eight cases alone. Prove-it™ Herpes revealed also individual targets not suspected initially by patient’s profile such as HHV-6, EBV and CMV. The final sensitivity and specificity for Prove-it™ Herpes assay were 90% and 98%. These results, especially HHV-7 cases, yield also interesting data for further investigations. Prove-it™ Herpes was considered as a rapid and robust diagnostic platform that was easily implemented into laboratory workflow. The broad target coverage and small sample volume required by the assay could benefit diagnostics, and thus, the treatment of life-threatening infections of the central nervous system.
PX-28 A comparison study of the biomerieux nucelisens easyMAG and the QIAGEN QIAsymphony extractors S.J. Shepherd1 , R.N. Gunson1 , T. Hanselle2 , J. Buckels3 , W.F. Carman1 *. 1 West of Scotland Specialist Virology Centre, Gartnavel General Hospital, Glasgow, Scotland, 2 QIAGEN GmbH, Hilden, Germany, 3 QIAGEN Ltd, Crawley, UK The West of Scotland Specialist Virology Centre (WOSSVC) is equipped with a variety of automated nucleic acid extraction machines including the BioMerieux easyMAG and the recently marketed QIAsymphony. Both machines have been described by their manufacturers as compact and easy to use. A direct comparison between the easyMAG and the QIAsymphony was undertaken to determine the suitability of the QIAsymphony to work in conjunction with and/or replace the easyMAG. A total of 300 (149 postitive/151 negative) respiratory, blood, tissue and swab samples were extracted simultaneously on both the easyMAG and the QIAsymphony. The eluted samples from both extractors were then tested in duplicate by qualitative real-time PCR and the cycle threshold (Ct) values for each extract compared. Overall the QIAsymphony performed similarly to the easyMAG, irrespective of sample type or pathogen listed. A small number of differences were seen between some samples. In most cases this was likely to be was related to the low level of virus present within the sample material. The main advantages of the QIAsymphony over the easyMAG are not only the benefit of extracting a larger number of samples per run but also the automated addition and elution of samples, the easyMAG requires both these steps to be carried out manually. The main disadvantage found with the QIAsymphony was the requirement for a specific set volume of starting sample, where the easyMAG can use volumes <200ml, the virus/bacteria kit of the QIAsymphony requires a volume in excess of 200ml.