Identification of RNA linkage isomers by anion exchange purification with electrospray ionization mass spectrometry of automatically desalted phosphodiesterase-II digests

Purification and characterisation of dsRNA using ion pair reverse phase chromatography and mass spectrometry

“…Mass spectrometry has emerged as an increasingly powerful tool for the identification and characterisation of nucleic acids. Prior to mass spectrometry analysis, the RNA of interest is routinely first purified using HPLC [31] , [32] , [33] , [34] , [35] . This often large biomolecule is subsequently digested with endonucleases into smaller oligoribonucleotides that are more amenable for chromatographic separation and intact mass measurements [36] , [37] .…”

Comparing ion‐pairing reagents and sample dissolution solvents for ion‐pairing reversed‐phase liquid chromatography/electrospray ionization mass spectrometry analysis of oligonucleotides

“…The emergence of soft ionization techniques for mass spectrometry, namely electrospray ionization (ESI) and matrix‐assisted laser desorption/ionization (MALDI), has substantially benefited the characterization of oligonucleotides by mass spectrometry (MS). However, mass measurement of oligonucleotides is complicated by the affinity of the polyanionic backbone to various cations including sodium and potassium which are often present in analyte solutions .…”

“…Therefore, quality control is a critical step in their synthesis and is usually accomplished by chromatographic techniques such as high-performance liquid chromatography (HPLC) [6][7][8][9][10] and electrophoresis. [11][12][13][14][15] The emergence of soft ionization techniques for mass spectrometry, namely electrospray ionization (ESI) and matrixassisted laser desorption/ionization (MALDI), [16][17][18][19][20][21] has substantially benefited the characterization of oligonucleotides by mass spectrometry (MS). However, mass measurement of oligonucleotides is complicated by the affinity of the polyanionic backbone to various cations including sodium and potassium which are often present in analyte solutions.…”

Mass Spectrometry of siRNA

“…A fraction-collecting autosampler was configured with a dual column selection valve using anion-exchange oligonucleotide purification as the first step followed by ion-pair reversed-phase desalting. The collected purified and desalted fractions were then injected into the mass spectrometer with a flow injection system to produce multiply charged species with minimal cation adducts [74]. Although purification and desalting examples were demonstrated with siRNA products from an enzymatic digestion reaction, the method should be applicable to analyzing siRNA from complex biological matrices.…”