Potential Primary Measurement Tool for the Quantification of DNA

Abstract

An automated sample introduction system, utilizing a demountable direct injection high-efficiency nebulizer (d-DIHEN), is successfully incorporated for the first time with an inductively coupled plasma optical emission spectrometer (ICP-OES) for the measurement of the phosphorus content in acid-digested nucleotides and deoxyribonucleic acid (DNA). With this experimental setup, the solution uptake rate and volume are reduced from 170 to 30 microL min(-1) and from 10 to 2.4 mL, respectively, thereby reducing the required DNA sample mass for solutions containing 3 microg g(-1) P from 300 to 72 microg of DNA, in comparison to previous analyses in our laboratory using a glass concentric nebulizer with cyclonic spray chamber arrangement. The use of direct injection also improves P (I) 213.617 nm sensitivity by a factor of 4 on average. A high-performance (HP) methodology in combination with the previous sample introduction system and ICP-OES provides simultaneous, time-correlated internal standardization and drift correction resulting in relative expanded uncertainties (% U) for the P mass fractions in the range of 0.1-0.4 (95% confidence level) for most of the thymidine 5'-monophosphate (TMP), calf thymus DNA (CTDNA), and plasmid DNA (PLDNA) analyses. The d-DIHEN with HP-ICP-OES methodology allows for the quantification of DNA mass at P mass fractions as low as 0.5 microg g(-1), further reducing the required DNA mass to 12 microg, with small uncertainty (< or = 0.4%). This successful approach will aid in the development and certification of nucleic acid certified reference materials (CRMs), particularly for these samples that are typically limited in volume.