Characterization and performance of injection molded poly(methylmethacrylate) microchips for capillary electrophoresis

Injection molded poly(methylmethacrylate) (IM-PMMA), chips were evaluated as potential candidates for capillary electrophoresis disposable chip applications. Mass production and usage of plastic microchips depends on chip-to-chip reproducibility and on analysis accuracy. Several important properties of IM-PMMA chips were considered: fabrication quality evaluated by environmental scanning electron microscope imaging, surface quality measurements, selected thermal/electrical properties as indicated by measurement of the current versus applied voltage (I-V) characteristic and the influence of channel surface treatments. Electroosmotic flow was also evaluated for untreated and O2 reactive ion etching (RIE) treated surface microchips. The performance characteristics of single lane plastic microchip capillary electrophoresis (MCE) separations were evaluated using a mixture of two dyes-fluorescein (FL) and fluorescein isothiocyanate (FITC). To overcome non-wettability of the native IM-PMMA surface, a modifier, polyethylene oxide was added to the buffer as a dynamic coating. Chip performance reproducibility was studied for chips with and without surface modification via the process of RIE with O2 and by varying the hole position for the reservoir in the cover plate or on the pattern side of the chip. Additionally, the importance of reconditioning steps to achieve optimal performance reproducibility was also examined. It was found that more reproducible quantitative results were obtained when normalized values of migration time, peak area and peak height of FL and FITC were used instead of actual measured parameters.     

On-line sample preconcentration by sweeping with dodecyltrimethylammonium bromide in capillary zone electrophoresis

On-line sample preconcentration of oligonucleotides with a new sweeping carrier was developed by using dodecyltrimethylammonium bromide (DTAB) below the critical micelle concentration (CMC). The sweeping results with DTAB below and above the CMC were compared. The use of DTAB below the CMC benefits the preconcentration of the oligonucleotides, while the use of DTAB above the CMC is good for hydrophobic small molecules. The factors affecting the sweeping results were optimized and this method was evaluated by constructing calibration curves for thrombin aptamers. The sweeping scheme produced a 112-fold sensitivity enhancement for the oligonucleotides relative to that run in a running buffer without DTAB. The sweeping method developed here can be a good reinforcement of the preconcentration scheme by sweeping when less-hydrophobic analytes or large negatively-charged molecules need to be preconcentrated.     

Fragmentation pathway studies of oligonucleotides in matrix-assisted laser desorption/ionization mass spectrometry by charge tagging and H/D exchange

The desorption and decompositions of synthesized oligonucleotides bearing fixed charge sites have been investigated by linear, delayed-extraction, reflecting and post-source decay mode matrix-assisted laser desorption/ionization (MALDI) mass spectrometry. In contrast to the conventional [M + H]⁺ forms of unmodified molecules where a proton is likely attached to a nucleobase, here the charge is fixed at one of the termini. In this case the observed fragment ions always incorporate the charge-tag. H/D exchange experiments provide no evidence for intramolecular migration of protons on the phosphate backbone to initiate the fragmentation event. New unique pathways of proton migration from the ribose have been elucidated and are rationalized by a charge-remote fragmentation pathway.