Robotic automation performs a nested RT-PCR analysis for HCV without introducing sample contamination.
The Polymerase Chain Reaction (PCR) is a popular method to amplify and detect specific RNA and DNA sequences. To obtain maximum performance of PCR, it is best performed by highly skilled technologists because of the complexity of the assay and the potential for laboratory contamination from the amplification products produced. We chose to automate this nested RT-PCR for hepatitis C assay to significantly reduce the need for manual pipetting while preserving the excellent non-contamination performance of the corresponding manual test. A three axis cartesian robotic pipetting station was equipped to perform RT-PCR using an on-board automated thermal cycling device. 104 sera were analyzed using this modified pipetting station and we found a very close agreement (100% sensitivity and 98% specificity) with results previously obtained by corresponding manual RT-PCR analysis. This study demonstrated a user-programmed robotic pipetting system could successfully automate a complex PCR assay without contamination. Our results suggest that use of robotic pipetting station can provide cost efficient alternative to performance of molecular diagnostic assays while demonstrating minimal inter sample contamination.
Mifflin
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TE
Department of Biomedical Engineering, University of Virginia Health Sciences Center, Charlottesville 22908, USA. tem6h@virginia.edu
Estey
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CA
Felder
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RA
D004587:Electrophoresis, Agar Gel / Q000379:methods; D016174:Hepacivirus / Q000235:genetics*; D012367:RNA, Viral / Q000032:analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000295:instrumentation / Q000379:methods*; D012371:Robotics; D012984:Software
D012367:RNA, Viral