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PCR VALIDATION TESTING UTILIZING A NOVEL DYNAMIC MEASUREMENT SYSTEM.
Sharon Adams , Christina Russ , Marcela Uribe , Francesco Marincola MD and David Stroncek MD . 1 DTM-HLA Laboratory, National Institutes of Health, Bethesda, MD, USA .
When PCR validation is done correctly it provides measurements which can allow for the assessment of accuracy and uniformity of the PCR block. Not only does it provide useful information pertinent to the end product, it is a necessity for accreditation. Most methods employ static temperature collection with a single probe. Generally, single probe systems allow for an average of 90sec to achieve the target temperature. This does not allow for an acquisition of temperature as it would occur during normal cycling parameters. Most single probe systems only allow for collection of data at pre-defined points of 95
C/40C. Our laboratory set out to find a temperature acquisition system which would provide temperatures in a dynamic environment. System criteria should include multi-probe head, ease of use, reproducibility of data, ability to follow data trends and rapid data collection. The Driftcon
system was utilized for purposes of this study. Data was collected from 18 PE thermocyclers using a standard run protocol which targeted temperatures 30C/95C/90C/50C/70C/60C. Average run time for this protocol was 20 minutes. Reports are automatically generated by the system which include results on accuracy and spread (uniformity). All results are compared with a global market database which allows for more accurate assessment of market specifications. 8/18 demonstrated accuracy problems at one of the achieved target temperatures. 6/18 demonstrated accuracy problems at a target temperature not tested by a single probe system. The conclusion of this study was that the Driftcon system yielded information which can help determine the efficacy of a thermocycler beyond data which is collected from a single probe system.