Описание: Oligonucleotides synthesized using phosphoramidite chemistry are typically analyzed and purified using ion-pair reversed-phase liquid chromatography (IP‐RPLC) and anion-exchange chromatography. The high salt content of anion‐exchange purification fractions impairs the ability of oligonucleotides to participate in ion pairing during IP-RPLC analysis. This leads to the need for sample desalting prior to IP-RPLC analysis, which is typically performed manually using centrifugal filters.1 In addition to suppressing the ion pairing of the oligonucleotides, the highly salty mobile phase in IEX separations is not compatible with mass spectrometry. This is also true for a variety of other bioseparations, such size exclusion and affinity. An investigator has the option of manually collecting the peaks, desalting offline and reinjecting them onto an LC/MS system. With the introduction of 2DLC control into Masshunter 11 software, 2D-LC/MS may be the simplest way accomplish the goal of mass measurement from salty first dimension separations. To accomplish this, we can use either SEC or IPRP in the second dimension. In this setup, the application of 2D-LC increases the workflow speed and avoids manual sample preparation.
Join Agilent Technologies LC Applications Scientist, Patrick Cronan, for an in-depth discussion of 2D LC/MS separation of oligonucleotides with full 2D control in Masshunter 11.
1. Krieger S.; Dickhut C. Agilent Technologies, Direct Analysis of In Process Oligonucleotides without Manual Purification, Application Note 2018
3 Key Learning Objectives:
- Understand how to couple multiple LC methods for oligonucleotide detection into one seamless application
- Learn the new technology included in Agilent’s 2DLC Software for Masshunter 11
- Make better transitions from 1D salty buffers to a 2nd dimension LCMS separation
Лектор: Patrick Cronan, LC Applications Scientist, Agilent Technologies, Inc.