Polymer solution-filled column for the analysis of antisense phosphorothioates by capillary electrophoresis

Shu Hui Chen, Ruey Tay Tzeng

Research output: Contribution to journalArticlepeer-review

15 Citations (Scopus)

Abstract

A capillary electrophoresis (CE) column filled with 13% poly(ethylene glycol) (PEG) solution is demonstrated to resolve different lengths of antisense phosphorothioates in 100 mu Tris-borate (pH 9.0) buffer containing 30% formamide at 50°C. Two sets of mixtures composed of 15-20 mars of either antisense phosphorothioate or phosphodiester oligonucleotides were synthesized based on a sequence of the antisense orientation directed against DNA-methyltransferase (denoted as MT-AS) and were used as model compounds. It was found that column coating reduced electroosmotic flow, as well as wall adsorption, and led to the separation of both phosphorothioate and phosphodiester molecules. Substantial peak broadening, however, specifically occurred to the phosphorothioates and was reduced statisfactorily by the addition of formamide into the buffer solution, raising the temperature, and raising the pH value. Under experimental conditions, a linear relationship between the migration time and the base number was observed, indicating that no peak compression artifacts existed. Without tedious pretreatment, antisense phosphorothioares were spiked into human serum, followed by water dilution, and then directly injected into the column. Separation of different lengths of phosphorothioates was observed using pressure injection, which did not suffer from injection bias.

Original languageEnglish
Pages (from-to)547-554
Number of pages8
JournalElectrophoresis
Volume20
Issue number3
DOIs
Publication statusPublished - 1999 Jan 1

All Science Journal Classification (ASJC) codes

  • Analytical Chemistry
  • Biochemistry
  • Clinical Biochemistry

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