Method development for genomic Legionella pneumophila DNA quantification by inductively coupled plasma mass spectrometry

摘要

The development of a method for the quantification of Legionella pneumophila genomic deoxyribonucleic acid is considered. The method is based on the quantification by inductively coupled plasma mass spectrometry (ICP-MS) of the mass fraction of phosphorus, stoichiometrically presented in the DNA molecules. Through the DNA sequencing data, it was possible to convert the ICP-MS analysis results into DNA genome units. L. pneumophila DNA samples were analyzed using ICP-sector field MS and ICP-quadrupole MS with a collision/reaction cell. Spectrophotometric measurements of the absorbance at 260 nm and real-time PCR techniques were used to independently confirm the ICP-MS results. The comparison of the methods showed that the ICP-MS method provides better accuracy with respect to currently applied analytical techniques such as UV spectrophotometry, fluorescent dye methods, and real-time PCR. Moreover, with the use of calibration standards whose values are traceable to the International System of Units and the possibility of evaluating the contribution to the overall uncertainty of each step of the measurement procedure, the method enables long-term comparability of the measurement results. These advantages make the ICP-MS method suitable for nucleic acid investigation, from nucleotides to genomic DNA, as well as for the certification of the reference materials containing nucleic acids.