(623bn) Developing a Miniaturized Assay for Label-Free, High-Throughput Biomolecule Analysis

Microfluidics technology has been widely applied to the miniaturization of analytical methods and chemical and biological processes because it has many advantages, such as significant reduction in analysis time, much lower waste production, and enhanced system performance and functionality by integrating different components onto microfluidic devices. These applications are usually called micro total analysis systems (µTAS) or lab on a chip (LOC). Among all the detection methods developed for microfluidic devices, laser-induced fluorescence (LIF) still remains the most popular because of its high sensitivity and compatibility with microfluidic devices. However, conventional fluorescence-based detection requires the analytes to be self-fluorescent or to be labeled with a fluorescence tag and thus results in limited applicability. Another issue of fluorescence-based detection is the analyte labeling process, which is not always straightforward and may cause undesired results, especially for biomolecules like proteins. UV absorbance-based detection has been a good alternative because it’s a label-free, nondestructive technique and has been widely used in many commercially available capillary electrophoresis and chromatography systems. However, the integration of UV absorbance-based detection into microfluidic devices is not always straightforward due to the short optical pathlength (usually between 5 and 50 µm) of typical microchannel designs. Additional difficulty comes from the need of UV-transparent material, usually quartz or fused silica, to construct microfluidic devices compatible with UV absorbance-based detection. This results in a significant increase in the cost and complexity of the device fabrication process.

To address these problems, we have designed and fabricated a new high-aspect-ratio microfluidic chip using a commercially available UV-curable glue and simple soft lithographic technique, and integrated it with a two-dimensional UV-absorbance imaging detector. The UV imaging detection technique we use here allows simultaneous detection of analytes in multiple microchannels on a single microchip with a high level of spatial and temporal resolution. As a proof of concept, we used this system to perform microchip DNA gel electrophoresis without labeling. This system can be extended to other biomolecule assays by changing the chip design. After optimization, we will use this system to perform a miniaturized, label-free protein immunoblot to validate the sexually dimorphic expression of two epigenetic protein biomarkers, H3K9/14Ac and H3K9Me3, from neonatal mouse brains.