Temperature-modulated array high-performance liquid chromatography

Using novel monolithic poly(styrene-divinylbenzene) capillary columns with ail internal diameter of 0.2 mm, we demonstrate for the first time the feas ibility of constructing high-performance liquid chromatography arrays for t he detection of mutations by heteroduplex analysis under partially denaturi ng conditions. In one embodiment, such an array can be used to analyze one sample simultaneously at different temperatures to maximize the detection o f mutations in DNA fragments containing multiple discrete melting domains. Alternatively, one may inject different samples onto columns kept at the sa me effective temperature. Further improvements in throughput can be obtaine d by means of laser-induced fluorescence detection and the differential lab eling of samples with up to four different fluorophores. Major advantages o f monolithic capillary high-performance liquid chromatographic arrays over their capillary electrophoretic analogs are the chemical inertness of the p oly(styrene-divinylbenzene) stationary phase, the physical robustness of th e Column bed due to its covalent linkage to the inner surface of the fused silica capillary, and the feasibility to modify the stationary phase thereb y allowing the separation of compounds not only on the principle of size ex clusion, but also adsorption, distribution, and ion exchange. Analyses time s are on the order of a few Minutes and turnaround time is extremely short as there is no need for the replenishment of the separation matrix between runs.