Elimination of false-positive polymerase chain reaction results resulting from hole punch carryover contamination

Journal of Veterinary Diagnostic Investigation : Official Publication of the American Association of Veterinary Laboratory Diagnosticians, Inc
Nicolai BonneS R Raidal

Abstract

The collection of biological material (e.g., blood) directly onto filter paper for subsequent use in laboratory assays such as polymerase chain reaction (PCR), has become a common practice. Dried cells or fluid on the paper can be readily rehydrated and retrieved into a standard volume of an appropriate elution buffer but introduces a dilution factor to the sample. The use of a common cutting instrument for excising a standard-sized piece of paper that contains the material also introduces the potential for transferring biological material from one sample to subsequent samples, causing false-positive results by PCR. In the present study, filter-paper-collected blood that contained beak and feather disease virus was used to determine if viral DNA could be transferred between samples by a hole punch used to excise sequential filter papers. It was determined that false-positive results could be obtained at least 13 times after a positive sample. Subsequently, the efficacy of 4 methods of hole punch disinfection, flaming, VirkonS, bleach, and a bleach-ethanol combination, was assessed. The only effective and practical method to destroy DNA was a method where the hole punch was agitated in commercial bleach, rinsed in water, the wat...Continue Reading

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Citations

Nov 17, 2011·Journal of Microbiological Methods·Christina A KelloggGary L Andersen
Oct 12, 2012·Journal of Clinical Microbiology·Sean C MurphyRobert Coombs
Dec 29, 2012·PLoS Neglected Tropical Diseases·Letícia Helena dos Santos MarquesMariângela Carneiro
Aug 13, 2013·PLoS Neglected Tropical Diseases·Soa Fy AndriamandimbyVincent Richard
Nov 26, 2011·Journal of Crohn's & Colitis·Herbert J Van Kruiningen
Aug 8, 2014·Journal of Virological Methods·Subir SarkerShane R Raidal
Apr 11, 2013·Journal of Clinical Virology : the Official Publication of the Pan American Society for Clinical Virology·Felix KleshikThomas R Battersby
Apr 14, 2015·Future Microbiology·Alison Luce-FedrowAllen L Richards
May 31, 2014·British Poultry Science·G SuriyapholS Dhitavat
May 12, 2009·Veterinary Pathology·M VaranatE B Breitschwerdt
Oct 15, 2013·Electrophoresis·Gi Won ShinGyoo Yeol Jung
Aug 30, 2019·Virus Genes·Shubhagata DasShane R Raidal

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Methods Mentioned

BETA
genotyping
PCR

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