Determination of therapeutic oligonucleotides using capillary gel electrophoresis

Oligonucleotides have developed into highly versatile and selective therapeutics over the past 20 years. More than five discrete mechanisms of action have been reported and more than 10 different chemical modifications have been used to extend their in vivo half-life and reduce their toxicity. Capillary gel electrophoresis (CGE) has been used extensively for the quantitative analysis of oligonucleotide therapeutics in both preclinical and clinical studies since the 1990s. The success of CGE is based on its extraordinary resolving power, which allows for the simultaneous determination of the parent drug and its metabolites. More recently, capillary gel electrophoresis has seen renewed interest with the emergence of replaceable gels with single-base resolving power and new capillary electrophoresis-mass spectrometry interfaces. This review discusses the bioanalysis of therapeutic oligonucleotides showing the evolution of the field over the past two decades leading to the current new approaches. Included in this review are topics such as different gel types, sample introduction modes, sample extraction procedures, separation conditions and detection methods used in CGE, along with discussions of the successes and limitations associated with each.