Nucleic acid biosensors

textThere is an ever increasing need to expand the realm of diagnostic capabilities. The need for accurate and reliable biological detection methods is crucial for genetic analysis, drug discovery, the monitoring of bacteria and viruses in food and water samples, as well as monitoring the environment for biothreat reagents. In the realm of diagnostics, molecular biology first expanded to include semiconductor manufacturing technology in the creation of a DNA biosensor array. Currently, biosensor arrays are being developed to utilizing complex sensors for the detection of multiple types of analytes. Important to the future applications for biosensor technology is the development of miniaturized systems that utilize sophisticated sensors. We have explored the use of platforms that offer a three-dimensional volume element, as well as sophisticated nucleic acid sensors. A flow cell based microsphere platform (electronic tongue), and a novel hydrogel platform based on shape recognition elements (MUFFINS), have been combined with nucleic acid sensors (ssDNA and aptamers) and fluorescence microscopy to create biosensors for the detection of DNA and proteins. As a step towards fulfilling the goal of developing a second generation of sophisticated sensors, signaling aptamers were engineered. Signaling aptamers are nucleic acid ligands (aptamers) that are capable of displaying changes in fluorescent signal upon binding to their cognate analyte. In addition, “affinity” microspheres were developed for the direct synthesis of microsphere bound nucleic acid sensors.Chemistr

Construction and selection of bead-bound combinatorial oligonucleoside phosphorothioate and phosphorodithioate aptamer libraries designed for rapid PCR-based sequencing

Chemically synthesized combinatorial libraries of unmodified or modified nucleic acids have not previously been used in methods to rapidly select oligonucleotides binding to target biomolecules such as proteins. Phosphorothioate oligonucleotides (S-ODNs) or phosphorodithioate oligonucleotides (S(2)-ODNs) with sulfurs replacing one or both of the non-bridging phosphate oxygens bind to proteins more tightly than unmodified oligonucleotides and have the potential to be used as diagnostic reagents and therapeutics. We have applied a split synthesis methodology to create one-bead one-S-ODN and one-bead one-S(2)-ODN libraries. Binding and selection of specific beads to the transcription factor NF-κB p50/p50 protein were demonstrated. Sequencing both the nucleic acid bases and the positions of any 3′-O-thioate/dithioate linkages was carried out by using a novel PCR-based identification tag of the selected beads. This approach allows us to rapidly and conveniently identify S-ODNs or S(2)-ODNs that bind to proteins

Analysis of Outcomes in Ischemic vs Nonischemic Cardiomyopathy in Patients With Atrial Fibrillation A Report From the GARFIELD-AF Registry

IMPORTANCE Congestive heart failure (CHF) is commonly associated with nonvalvular atrial fibrillation (AF), and their combination may affect treatment strategies and outcomes