Modification of poly(methyl methacrylate) microchannels for highly efficient and reproducible electrophoretic separations of double-stranded DNA

This paper deals with dynamic coating of the microchannels fabricated on poly(methyl methacrylate) (PMMA) chips and DNA separation by microchip electrophoresis (MCE). After testing a number of polymers, including 2-hydroxyethyl cellulose, hydroxypropylmethyl cellulose, different sizes of poly(ethylene oxide) (PEO), and poly(vinyl pyrrolidone) (PVP), we found that coating of the PMMA microchannels with PEO(Mr = 6.0 × 105 g/mol) on the first layer is essential to minimize the interaction of DNA with PMMA surface. To achieve high efficiency, multilayer coating of PMMA chips with PEO, PVP, and PEO containing gold nanoparticles [PEO(GNP)] is important. A 2-(PEO-PVP)-PEO(GNP) PMMA chip, which was repeatedly coated with 1.0% PEO and 5.0% PVP twice, and then coated with 0.75% PEO(GNP) each for 30 min, provided a high efficiency (up to 1.7 × 106 plates/m) for the separation of DNA markers V (pBR 322/HaeIII digest) and VI (pBR 328/BglI digest and pBR 328/HinfI digest) when using 0.75% PEO(GNP). With such a high efficiency, we demonstrated the separation of hsp65 gene fragments of Mycobacterium HaeIII digests by MCE within 90 s. The advantages of this approach to DNA analysis include ease of filling the microchannel with 0.75% PEO(GNP), rapidity, and reproducibility.