A Novel Microfluidic Approach for Denaturating High-resolution DNA Electrophoresis

RP-35 The Agilent 2100 Bioanalyzer instrument platform is designed for performing fast electrophoretic analysis of DNA, RNA and proteins based on microfluidic chip technology. It is widely used for the analysis of PCR products and restriction digests. However, the resolution of the current DNA assays is not sufficient to separate DNA fragments generated in applications like e.g. STR-typing, AFLP and MLPA. As these methods require a very high resolving power in the electrophoresis involving denaturing conditions, usually capillary-based DNA sequencers are used to separate the DNA fragments. However, due to a growing number of new applications, there is a strong demand for an alternative separation platform that is fast, flexible and easy to use. Here we describe a novel Bioanalyzer assay prototype that is able to perform fast, sensitive, high resolution separation of labeled DNA fragments on a modified Agilent 2100 Bioanalyzer instrument. A new denaturating gel matrix has been developed that enables the separation of DNA fragments between 10 and 550 bp with a resolution approaching 2 basepairs. By the use of an internal size standard sizing precision results with standard deviations of 0.1 to 0.2 bp were achieved. The significantly increased resolution of the new assay together with the possibility to use an internal size standard enables applications like STR-typing and MLPA analysis on the Bioanalyzer.