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Evaluation of real-time PCR based assays for the detection and quantitation of CMV DNA

Abstract number: 1733_946

Rudolph M., Willert B., Grewing T., Fuhrmann J.

Introduction: Infections with human cytomegalovirus (CMV) are usually asymptomatic or related with harmless symptoms. But severe problems occur if embryos, transplant patients, patients under immunosuppressive therapy or AIDS patients get infected. CMV infection in this risk group can even be fatal. The aim of this work was the evaluation of real-time PCR assays for the fast, sensitive and specific detection of CMV DNA and the reliable quantitation of the viral load. These assays are important and useful for testing organ donors, monitoring of CMV patients under antiviral therapy, and for the protection of unborn children.

Materials and Methods: The artus® PCR Kits were designed and optimised for different real-time PCR instruments: LightCycler®, the Rotor Gene™ 3000 and the ABI PRISM®7000–7900 Instruments. The analytical sensitivity was determined by probit analysis. Using negative human EDTA-plasma was spiked with pre-quantified CMV. Serial dilutions were prepared, purified using the QIAamp® DSP Virus Kit and analysed with the respective artus CMV PCR Kit. The analytical specificity was tested with isolates of related viruses and other human pathogens. The diagnostic specificity and sensitivity was determined by analysing 177 samples in comparison with the COBAS AMPLICOR™ CMV MONITOR System.

Results: The determined analytical sensitivity was 65, 57 and 64 copies/mL for the LightCycler Instrument, the Rotor-Gene 3000, and the ABI PRISM assay, respectively. Cross reactivity was excluded testing isolates of other pathogens. The diagnostic sensitivity and specificity of the artus CMV PCR Kits was determined by comparison with the COBAS AMPLICOR CMV MONITOR System. Clinical sensitivity of 100% was determined.

Conclusions: The results demonstrate that the artus CMV PCR Kits, fulfil all requirements for a fast, sensitive, and specific detection of CMV DNA and the reliable quantitation of the CMV viral load. Therefore, the artus CMV PCR Kits can be perfectly used for the successful management of CMV disease, especially in immunocompromised patients and pregnant women.

Trademarks: QIAGEN®, artus®, QIAamp®, (QIAGEN Group), ABI PRISM® (Applera Corporation), CORBAS AMPLICORTM, LightCycler® Instruments (Roche Group), Rotor-GeneTM (Corbett Research)

Session Details

Date: 31/03/2007
Time: 00:00-00:00
Session name: European Society of Clinical Microbiology and Infectious Diseases
Subject:
Location: ICC, Munich, Germany
Presentation type:
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