Photo-polymerized cross-linked polyacrylamide gels are becoming increasingly important for use in micro-fabricated DNA electrophoresis systems because they allow a concentrated sieving matrix to be precisely positioned at any location within a complex micro-channel network. The rate of photo-initiation in the free radical gel polymerization reaction, however, can exert a strong influence on the resulting gel structure. Experimental data on separation resolution of single stranded DNA (ssDNA) in photoinitiated polyacrylamide gels is very sparse. In this study, we investigate the performance of ssDNA electrophoresis in an ALF Express automated DNA sequencer using various photo-initiation chemistries. Cross-linked polyacrylamide gels with concentrations ranging from 6 to 12 %T were prepared using riboflavin, methylene blue, irgacure 651 and ReproGel (AP Biotech) photo-initiators. Separation resolution is compared with that attained in cross-linked polyacrylamide gels prepared using conventional chemical initiators (e.g. ammonium persulfate/ Tetramethylethylenediamine) in order to determine the polymerization conditions necessary for optimum performance.

Spider-silk is known as one of the stronger natural materials, unfortunately it is impossible to farm spiders due to their territorial and cannibalistic nature. To address this issue, researchers have studied spider-silk to discover how it is produced in nature. From their results, spider-silk is composed of large sized proteins produced in two different cell types. Using this knowledge, researchers created transgenic organisms capable of producing spider-silk proteins in large quantities. Using these proteins, several groups have created fibers, films, hydrogels, and adhesives with robust and versatile properties. Wet-spinning is a technique commonly used to create fibers from spider-silk proteins. These fibers unfortunately do not compare to the mechanical properties of natural silk. To address this researchers have used a method known as electrospinning to create spider-silk fibers with substantially smaller diameters. In doing so, these electrospun fibers have increased surface area and enhanced mechanical properties. Using this method, our group has modified the electrospinner to be able to produce continuous fine diameter yarns composed of hundreds of nanofibers with mechanical properties surpassing that of natural silk. Fibers aside, spider-silk proteins can be used to create a variety of different biocompatible materials. To further enhance these materials, our group has utilized a technique traditionally used for observation. This technique employs a high intensity light source to initiate cross-links within the proteins. With this method, our spider-silk protein materials have increased their mechanical properties by a factor of seven. These materials can further be modified through post-treatments, resulting in tunable materials with diverse and robust mechanical properties.

There is great interest in miniaturized analytical systems for life science research, the clinical environment, drug discovery, biotechnology, quality control, and environmental monitoring and numerous articles have been written which predict the success of microfluidic based systems. It was demonstrated in this work that a microfluidic flow system could be quickly and easily manufactured in a research lab environment without the need for clean room facilities. The microfluidic device was created using polymethylmethacrylate, a CO2 laser and a standard oven. The device was designed, manufactured and ready for use within three hours. This work also investigated a chemiluminescent system which was intended for use in protease assays in the microfluidic device. This work also focused on the use of photoinitiated polymer monoliths, with immobilized tannic acid, as protein preconcentrators. The function of the monolithic devices was demonstrated by pumping low concentration solutions of BSA BODIPY® FL through the monolith. Both loading and elution were done using pressure. It was shown that BSA could be concentrated on and successfully eluted from the monolith. The elution volume for a 125 nl monolith was found to be 4 μl. Therefore an injection of a 60 μl sample of 1 x 10⁻⁹M BSA BODIPY ® FL gave rise to a concentration factor of 15. The pH optimum for the binding of BSA BODIPY ® FL was found to be pH 8.0 and the loading capacity of the tannic acid monolith was found to be 0.6 mg.ml⁻¹.

ABSTRACT Mixed function oxygenase system participates in biosynthesis of endogenous and metabolism of exogenous substances (e.g. drugs or chemical procarcinogens) in an organism. Substrates are biotransformed by terminal oxygenases - cytochromes P450 (P450). Catalytic properties of certain P450s (e.g. studied isoform 2B4) are altered in the presence of a redox partner - cytochrome b5 (cyb5). Both cytochromes are anchored by hydrophobic domains in a lipid membrane of endoplasmic reticulum whereas their catalytic domains are exposed to cytosol. Two zero-length cross-linking approaches were employed to extend present knowledge of P450 2B4 and cyb5 protein structure and protein-protein interactions: (1) interlinking of carboxylate and primary amine groups of amino acids by water soluble 1- ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC), and (2) photo-initiated cross-linking by photo-labile methionine analog (pMet), which links to any amino acid after activation by UV-irradiation, either in hydrophilic or hydrophobic environment. pMet was incorporated to methionine site(s) of cyb5 during recombinant expression in E. coli, which was carried out in limit medium supplemented with amino acid analog. Optimization of experimental conditions led to ~20-30% substitution of the natural amino acid. Covalent...