Integrated Sample Cleanup-Capillary Electrophoresis Microchip for High-Performance Short Tandem Repeat Genetic Analysis
ABSTRACT An integrated PCR sample cleanup and preconcentration process is developed for forensic short tandem repeat (STR) analysis using a streptavidin-modified photopolymerized capture gel injector for microchip capillary electrophoresis (microCE). PCR samples generated with one biotinylated primer and one fluorescent primer provide the input to the streptavidin-based affinity capture-microCE device. Monoplex PCR samples processed by the device exhibited approximately 10- to 50-fold increased fluorescence intensities, and DNA profiles generated using 9-plex STR samples displayed approximately 14- to 19-fold higher signal intensities compared to those analyzed using traditional cross injection. Complete STR profiles were obtained with as few as 25 copies of DNA template using the capture-microCE device. Four DNA samples with various degrees of degradation were also tested. Samples analyzed using the capture-microCE device resulted in a significant increase of successful allele detection. The ability of our capture-microCE device and method to remove contaminating ions, to concentrate the sample injection plug, and to eliminate electrokinetic injection bias provides a powerful approach for integrating sample cleanup with DNA separation.
- SourceAvailable from: Augusto M Tentori
[Show abstract] [Hide abstract]
- "These advantages provide a 25-fold increase in separation efficiency yielding efficient multiplexed assays. Oligonucleotide capture regions have also been created using PA gels copolymerized with streptavidin (Yeung et al 2009). Subsequent to PA gel polymerization, biotinylated oligonucleotides were immobilized via streptavidin in the gel regions, thus avoiding the need to copolymerize oligonucleotides directly into the gel matrix. "
ABSTRACT: Microfluidic technologies are playing an increasingly important role in biological inquiry. Sophisticated approaches to the microanalysis of biological specimens rely, in part, on the fine fluid and material control offered by microtechnology, as well as a sufficient capacity for systems integration. A suite of techniques that utilize photopatterning of polymers on fluidic surfaces, within fluidic volumes, and as primary device structures underpins recent technological innovation in bioanalysis. Well-characterized photopatterning approaches enable previously fabricated or commercially fabricated devices to be customized by the user in a straight-forward manner, making the tools accessible to laboratories that do not focus on microfabrication technology innovation. In this review of recent advances, we summarize reported microfluidic devices with photopatterned structures and regions as platforms for a diverse set of biological measurements and assays.Journal of Micromechanics and Microengineering 05/2011; 21(5):54001. DOI:10.1088/0960-1317/21/5/054001 · 1.73 Impact Factor
- [Show abstract] [Hide abstract]
ABSTRACT: Functionalities of microchip-based analytical systems for nucleic acids analysis are highlighted and driving forces for the future progress are reviewed. The review shows the main advantages of microchip-based systems and discusses up-to-date achievements in this area. Among the main advantages are a high speed of analysis, high rates of heating and cooling, a low reagents consumption, and a high potential to integrate several analytical processes in a single microchip device.Russian Journal of General Chemistry 12/2013; 82(12). DOI:10.1134/S1070363212120353 · 0.48 Impact Factor
- Journal of Vascular and Interventional Radiology 02/2002; 13(2). DOI:10.1016/S1051-0443(02)70096-1 · 2.41 Impact Factor