This paper lays out the production, analytical methods, and statistical evaluations required for the production of the Standard Reference Material 2372a, Human DNA Quantitation Standard, including storage and use, background, and experimental methods; it includes four appendixes detailing DNA extraction method, spectrophotometric measurement protocol, SRM 2372a statistician’s report, and performance in commercial qPCR chemistries.
This paper documents the production, analytical methods, and statistical evaluations involved in the production of the Standard Reference Material (SRM) 2372a, Human DNA Quantitation Standard. The manual is intended for use in the value assignment of human genomic deoxyribonucleic acid (DNA) forensic quantitation materials. The three SRM 2372a components are human genomic DNA extracts prepared at the National Institute of Standards and Technology (NIST) from buffy coat white blood cells from single-source anonymous donors. SRM 2372a describes the storage, use, and history and background of DNA quantitation; experimental methods, including polymerase chain reaction (PCR) assays, droplet digital PCR measurements, sample preparation, absorbance measurements, and more experimentation methods; measurement results, including absorbance spectrophotometry, droplet volume, certification measurements, mitochondrial to nuclear DNA ratio (mtDNA/nDNA), and performance in commercial qPCR chemistries. The manual also reviews the qualification of ddPCR as fit-for-purpose, and provides conclusions and recommendations for certification values as well as use of this manual for DNA quantitation of SRM 2372a for value-assigning in-house reference materials. The document includes a linked table of contents, four appendixes, and a number of tables displaying examples and other relevant data and visual aids.