Systematic optimization of gel-free capillary zone electrophoresis for phosphodiester oligonucleotide separations at acidic pH
Year: 2026
Type: article
Abstract: BACKGROUND: Reliable analysis of oligonucleotides is essential for therapeutic development and biomarker studies, yet conventional methods often rely on gels or high-salt buffers that hinder mass spectrometry compatibility. MicroDNA and microRNA are commonly analyzed using liquid chromatography or capillary gel electrophoresis (CGE). In a recent study, we introduced a gel-free capillary zone electrophoresis (CZE) method for rapid, high-resolution separation of oligonucleotides at acidic pH. RESULTS: Here we optimized a gel-free capillary zone electrophoresis (CZE) method operating at acidic pH using a PVA-coated capillary and malonic acid-ammonia buffer. Adjustments to buffer composition, cation type, and temperature (up to 60 °C) enhanced resolution and reduced current, enabling rapid, high-resolution separation of phosphodiester oligonucleotides ranging from 18 to 25 nucleotides with excellent repeatability (<0.5% mobility variation). SIGNIFICANCE: This gel-free CZE approach provides a fast and reproducible platform, facilitating detailed characterization of phosphodiester oligonucleotides, microDNA, and microRNA in pharmaceutical and bioanalytical contexts.
Source: Analytica Chimica Acta
Institution: Uppsala University
Language: English
Subfield: Biomedical Engineering
Field: Engineering
Domain: Physical Sciences
Open Access status: hybrid
Estimated APC paid: $4,270
OpenAlex ID: W7135376339