Making standards for quantitative real-time pneumococcal PCR

Susan C Morpeth; Jim F Huggett; David R Murdoch; J Anthony G Scott

doi:10.1016/j.bdq.2014.11.003

Making standards for quantitative real-time pneumococcal PCR

Biomol Detect Quantif. 2014 Dec 4:2:1-3. doi: 10.1016/j.bdq.2014.11.003. eCollection 2014 Dec.

Authors

Susan C Morpeth¹, Jim F Huggett², David R Murdoch³, J Anthony G Scott⁴

Affiliations

¹ KEMRI-Wellcome Trust Research Programme, Kilifi, Kenya; Nuffield Department of Clinical Medicine, University of Oxford, Oxford, United Kingdom.
² Molecular & Cell Biology, LGC, London, United Kingdom.
³ Department of Pathology, University of Otago, Christchurch, New Zealand; Microbiology Unit, Canterbury Health Laboratories, Christchurch, New Zealand.
⁴ KEMRI-Wellcome Trust Research Programme, Kilifi, Kenya; Nuffield Department of Clinical Medicine, University of Oxford, Oxford, United Kingdom; Department of Infectious Disease Epidemiology, London School of Hygiene and Tropical Medicine, London, United Kingdom.

Abstract

Quantitative lytA PCR is often performed using in-house standards. We hypothesised equivalence when measuring a standard suspension of Streptococcus pneumoniae by colony-forming-units (CFU) or genome-copies. Median (IQR) ratio of CFU/genome-copies was 0.19 (0.1-1.2). Genome-copies were less variable than CFU, but the discrepancy between the methods highlights challenges with absolute quantification.

Keywords: Quantitative; Real-time PCR; Standards; Streptococcus pneumoniae.