HIGH-SPEED SEPARATION OF ANTISENSE OLIGONUCLEOTIDES ON A MICROMACHINED CAPILLARY ELECTROPHORESIS DEVICE

A micromachined chemical analysis device based on capillary electropho resis has been successfully used for very fast size separation of a sy nthetic mixture of fluorescent phosphoro-thioate oligonucleotides rang ing from 10 to 25 bases in length. The device consists of a channel sy stem which has been formed in the surface of a glass plate by a standa rd photolithographical process. An integrated volume-defined sample in jector allowed for unbiased electrokinetic introduction of sample plug s of 150-mu m length (corresponding sample volume, 90 pL) into the sep aration channel. This well-defined injection procedure, in combination with the application of high electric fields of up to 2300 V/cm, resu lted in size separation of single-stranded oligonucleotides in less th an 45 s when a separation distance of 3.8 cm was used. Column efficien cies of up to 200 000 theoretical plates with an associated plate heig ht of 0.2 mu m were obtained. Fast repetitive sample injection and ana lysis could be demonstrated with excellent reproducibilities for both migration time (<0.06%) and peak height (<1.7%). The results might als o be of relevance for DNA sequencing, where fast oligonucleotide analy sis is of key importance. Moreover, they provide a route to ''on-line' ' analysis of synthetic and natural oligonucleotides and possibly othe r classes of biopolymers.