Fully Automated Nucleic Acid Extraction: MagNA Pure LC

Abstract

Kinetic PCR analysis by real-time monitoring of DNA amplification was first described 8 years ago (1). Recently, the LightCyclerTM instrument (Roche Molecular Biochemicals, Mannheim, Germany), which allows high-speed thermal cycling by use of air instead of thermal blocks and on-line real-time fluorescence monitoring, was introduced (2). Several reports have been published with regard to utilization of the LightCycler technology for detection of pathogens (3)(4)(5)(6)(7)(8)(9)(10)(11). Real-time PCR has greatly decreased the amount of hands-on time needed to generate and detect amplification products. Before amplification, pathogen-specific DNA or RNA must be extracted from the specimen. This procedure, also called sample preparation, remains the most labor-intensive and time-consuming part of widely automated molecular assays and may be considered the major weakness in most molecular assays today (12)(13). Therefore, a fully automated sample preparation system is urgently needed for the routine diagnostic laboratory. In this study, a fully automated specimen preparation instrument, the MagNA Pure LCTM (Roche) was evaluated. The new instrument was used for extraction of herpes simplex virus (HSV) DNA in combination with real-time PCR on the LightCycler instrument. In the first experiment, the interassay variation and the detection limit were tested. The Second European Union Concerted Action HSV Proficiency Panel, which consists of 12 vials with different concentrations of HSV type 1 (HSV-1), strain MacIntyre (American Type Culture Collection), HSV type 2 (HSV-2), strain MS (American Type Culture Collection), varicella-zoster virus, and negative samples, was used. Samples were analyzed three times on different days. Results were compared with a molecular assay, which consisted of an in-house DNA extraction protocol and real-time PCR. In the second experiment, intraassay variation of the new molecular assay was tested. Plasma was collected from a …