In third generation sequencing, long DNA molecules of more than ∼20 kbp are needed to obtain quality sequence data. Here we report a versatile technology for DNA size selection that fulfills this requirement.

Electrohydrodynamic migration, which is based on hydrodynamic actuation with an opposing electrophoretic force, enables the separation of DNA molecules of 3 to 100 kbp in capillary systems within one hour. Here, we wish to enhance these performances using microchip technologies. This study starts with the development of a fabrication process to obtain microchips with uniform surfaces, which is motivated by our observation that band splitting occurs in microchannels made out of heterogeneous materials such as glass and silicon. The resulting glass-adhesive-glass microchips feature the highest reported bonding strength of 11 MPa for such materials (115 kgf/cm 2), a high lateral resolution of critical dimension 5 µm, and minimal auto-fluorescence. These devices enable us to report the separation of 13 DNA bands in the size range of 1-150 kbp in one experiment of 5 minutes, i.e. 13 times faster than with capillary electrophoresis. In turn, we provide evidence that band splitting in heterogeneous glasssilicon microchips arises from differential Electro-Osmotic Flow (EOF) in between the upper and lower walls of the channel. We further indicate that differential EOF occurs in microchip electrophoresis and constitutes an underappreciated source of band broadening. We finally prove that our separation data compare favorably to state-of-the-art microchip technologies in terms of resolution length and theoretical plate numbers.