Enhancing the Mass Spectrometry Sensitivity for Oligonucleotide Detection by Organic Vapor Assisted Electrospray

Abstract

There are two challenges in oligonucleotide detection by liquid chromatography coupled with mass spectrometry (LC-MS), the serious ion suppression effects caused by ion-pair reagents and the low detection sensitivity in positive mode MS. In this study, highly concentrated alcohol vapors were introduced into an enclosed electrospray ionization chamber, and oligonucleotides could be well detected in negative mode MS even with 100 mM triethylammonium acetate (TEAA) as an ion-pair reagent. The MS signal intensity was improved 600-fold (for standard oligonucleotide dT15) by the isopropanol vapor assisted electrospray, and effective ion-pair LC separation was feasibly coupled with high-sensitive MS detection. Then, oligonucleotides were successfully detected in positive mode MS with few adducts by propanoic acid vapor assisted electrospray. The signal intensity was enhanced more than 10-fold on average compared with adding acids into the electrospray solution. Finally, oligonucleotides and peptides or histones were simultaneously detected in MS with little interference with each other. Our strategy provides a useful alternative for investigating the biological functions of oligonucleotides.

Online porous graphic carbon chromatography coupled with tandem mass spectrometry for post‐translational modification analysis

“…However, it is worth noting that the sensitivity dropped approximately 4‐fold due to the ion‐suppression effects of TFA. It is anticipated that the development of new ionization techniques will minimize the ion suppression, such as using an organic vapor electrospray source that can remove TFA during the ESI process . Nevertheless, the result confirmed the difference in the separation mechanisms between PGC and RPLC, and the PGC/MS/MS system is an alternative to RP/MS/MS in the proteomics field.…”

Current State of Oligonucleotide Characterization Using Liquid Chromatography–Mass Spectrometry: Insight into Critical Issues

“…Improved understanding of ways to advantageously exploit these mechanisms is a significant challenge which must be addressed to continue to improve ionization efficiency and therefore method sensitivity. One possibility to increase sensitivity is increasing the amount of organic present, like with HILIC chromatography or organic vapor assisted electrospray . The organic is known to play an important role in the desorption of oligonucleotides by decreasing droplet surface tension via changing the organic solvent or increasing the concentration of organic present …”

“…One possibility to increase sensitivity is increasing the amount of organic present, like with HILIC chromatography or organic vapor assisted electrospray. 31 The organic is known to play an important role in the desorption of oligonucleotides by decreasing droplet surface tension via changing the organic solvent or increasing the concentration of organic present. 11…”

Oligonucleotides Isolation and Separation—A Review on Adsorbent Selection

“…Interestingly, Biba et al showed that porous layer thickness influences OGN separation [ 10 ]. The narrower peaks were obtained for the column packed with particles of a smaller porous layer (0.35 µm) compared to those with the 0.5 µm layer [ 16 ]. On the other hand, Gilar and Bouvier [ 11 ] compared two columns packed with 5 µm adsorbents with 100 and 300 Å pores.…”

“…C18 modified silica is frequently used in IPC to separate shortmers, namely in vivo and in vitro metabolites [ 11 , 13 , 16 , 39 , 40 , 41 , 42 , 43 , 44 , 45 , 46 , 48 ]. Dai et al [ 39 ] presented successful baseline separation of phosphorothioate ASOs and five metabolites shorter with one to five nucleotides from 3′-end.…”