PMID- 31638773
OWN - NLM
STAT- MEDLINE
DCOM- 20201103
LR  - 20210110
IS  - 1520-6882 (Electronic)
IS  - 0003-2700 (Print)
IS  - 0003-2700 (Linking)
VI  - 91
IP  - 22
DP  - 2019 Nov 19
TI  - Absolute Quantification of RNA or DNA Using Acid Hydrolysis and Mass 
      Spectrometry.
PG  - 14569-14576
LID - 10.1021/acs.analchem.9b03625 [doi]
AB  - Accurate, traceable quantification of ribonucleotide or deoxyribonucleotide 
      oligomers is achievable using acid hydrolysis and isotope dilution mass 
      spectrometry (ID-MS). In this work, formic acid hydrolysis is demonstrated to 
      generate stoichiometric release of nucleobases from intact oligonucleotides, 
      which then can be measured by ID-MS, facilitating true and precise absolute 
      quantification of RNA, short linearized DNA, or genomic DNA. Surrogate 
      nucleobases are quantified with a liquid chromatography-tandem mass spectrometry 
      (LC-MS/MS) workflow, using multiple reaction monitoring (MRM). Nucleobases were 
      chromatographically resolved using a novel cation-exchange separation, 
      incorporating a pH gradient. Trueness of this quantitative assay is estimated 
      from agreement among the surrogate nucleobases and by comparison to 
      concentrations provided for commercial materials or Standard Reference Materials 
      (SRMs) from the National Institute of Standards and Technology (NIST). Comparable 
      concentration estimates using NanoDrop spectrophotometry or established from 
      droplet-digital polymerase chain reaction (ddPCR) techniques agree well with the 
      results. Acid hydrolysis-ID-LC-MS/MS provides excellent quantitative selectivity 
      and accuracy while enabling traceability to mass unit. Additionally, this 
      approach can be uniquely useful for quantifying modified nucleobases or mixtures.
FAU - Lowenthal, Mark S
AU  - Lowenthal MS
AUID- ORCID: 0000-0001-5795-8824
AD  - Biomolecular Measurement Division , National Institute of Standards and 
      Technology , 100 Bureau Drive, Stop 8314 , Gaithersburg , Maryland 20899 , United 
      States.
FAU - Quittman, Eva
AU  - Quittman E
AD  - Biomolecular Measurement Division , National Institute of Standards and 
      Technology , 100 Bureau Drive, Stop 8314 , Gaithersburg , Maryland 20899 , United 
      States.
FAU - Phinney, Karen W
AU  - Phinney KW
AUID- ORCID: 0000-0001-7540-5860
AD  - Biomolecular Measurement Division , National Institute of Standards and 
      Technology , 100 Bureau Drive, Stop 8314 , Gaithersburg , Maryland 20899 , United 
      States.
LA  - eng
GR  - 9999-NIST/ImNIST/Intramural NIST DOC/United States
PT  - Journal Article
DEP - 20191101
PL  - United States
TA  - Anal Chem
JT  - Analytical chemistry
JID - 0370536
RN  - 0 (DNA, Viral)
RN  - 0 (Deoxyribonucleotides)
RN  - 0 (Formates)
RN  - 0 (Ribonucleotides)
RN  - 0YIW783RG1 (formic acid)
RN  - 63231-63-0 (RNA)
SB  - IM
MH  - BK Virus/chemistry
MH  - Chromatography, Liquid/*methods
MH  - DNA, Viral/*analysis/chemistry
MH  - Deoxyribonucleotides/analysis/chemistry
MH  - Formates/chemistry
MH  - Humans
MH  - Hydrolysis
MH  - RNA/*analysis/chemistry
MH  - Ribonucleotides/analysis/chemistry
MH  - Tandem Mass Spectrometry/*methods
PMC - PMC7337568
MID - NIHMS1588439
EDAT- 2019/10/23 06:00
MHDA- 2020/11/04 06:00
PMCR- 2020/11/19
CRDT- 2019/10/23 06:00
PHST- 2019/10/23 06:00 [pubmed]
PHST- 2020/11/04 06:00 [medline]
PHST- 2019/10/23 06:00 [entrez]
PHST- 2020/11/19 00:00 [pmc-release]
AID - 10.1021/acs.analchem.9b03625 [doi]
PST - ppublish
SO  - Anal Chem. 2019 Nov 19;91(22):14569-14576. doi: 10.1021/acs.analchem.9b03625. 
      Epub 2019 Nov 1.