Comprehensive Evaluation of Multiple Automated High Throughput Extraction Platforms Using qPCR Assays

Detection of nucleic acid by quantitative polymerase chain reaction (qPCR) and / or quantitative reverse transcriptase PCR (qRT-PCR) continues to be the gold standard in supporting biodistribution, vector shedding, gene expression, and pharmacokinetics bioanalysis studies. These molecular platforms are highly preferred due to their inherent sensitive and robust nature. Furthermore, qPCR and RT-qPCR assays continue to play a central role in clinical diagnostics wherein, clinicians heavily rely on these assays to diagnose (assess levels of viral and bacterial pathogens) and effectively treat patients. Analysis of viral nucleic acid in biological matrices requires efficient extraction of viral nucleic acids. Selection of an appropriate extraction platform is key to the successful method development / validation / testing of any qPCR / RT-qPCR assay. Selections are highly dependent on unique properties of biological matrices. Importantly, introducing automated extraction platforms into a laboratory’s workflow increases the efficiency and consistency of the test results. Automated magnetic and silica-based extraction technologies are commonly used in clinical diagnostics and bioanalysis laboratories. In this study, we describe the evaluation of 24 well NucliSENS®easyMag® (BioMerieux; magnetic bead based), 96 well KingFisherTM Flex (Thermo Fisher Scientific; magnetic bead based), 12 well QIAcube Connect (Qiagen; silica based), and 96 well QIAcube HT (Qiagen; silica based) using serum, neat urine, plasma, and peripheral blood mononuclear cells (PBMCs). We assessed detection of Epstein-Barr virus (EBV), Varicella zoster virus (VZV), Herpes simplex virus 1 (HSV-1), and Severe acute respiratory syndrome coronavirus 2 (SARS‑CoV‑2) in urine, serum, plasma, and PBMCs.