ang-Anh Ho

Optical Sensors Based on Hybrid Aptamer/Conjugated Polymer Complexes

Single-stranded DNA (aptamer) can specifically bind potassium ions or human alpha-thrombin. When binding takes place, the aptamer undergoes a conformational transition from an unfolded to a folded structure. This conformational change of the negatively charged oligonucleotide can be detected by adding a water-soluble, cationic polythiophene derivative, which transduces the new complex formation into an optical (colorimetric or fluorometric) signal without any labeling of the probe or of the target. This simple and rapid methodology has enabled the detection of human thrombin in the femtomole range. This new biophotonic tool can easily be applied to the detection of various other proteins as well as being useful in the high-throughput screening of new drugs.

Colorimetric and fluorometric detection of nucleic acids using cationic polythiophene derivatives.

Simple and reliable sequence-specific methods are needed for the rapid detection of oligonucleotides, to diagnose infections and various genetic diseases. In this regard, interesting optical and electrochemical DNA-hybridization sensors have been proposed.[1±5] The recognition capabilities of DNA are well established but, to transduce the recognition event into a physically measurable value, a fluorescent or electroactive tag is often bound to the analyte. Electrochemical and optical sensors based on conjugated polymers have also been reported[6±9] and some oligonucleotide-functionalized conjugated polymers can also transduce hybridization events into an electrical signal without labeling of the oligonucleotide target.[10±12] The detection relies on a modTo our knowledge this is the first report on the use of singlemolecule atomic-force spectroscopy to study the reduction pathway of multiple disulfide bonds in proteins and to evaluate the distributions of intermediates obtained under different reducing conditions without separating them and without any blocking and fractionation steps. The characterization of these intermediates has so far been accomplished by first blocking them with reagents such as alkylalkanethiosulfonates and then by fractionation by ion-exchange chromatography, 2D or capillary gel electrophoresis, or gel filtration.[11] The determination of thiol groups and disulfide bonds in a polythiol systems has always been a very challenging problem.[12] The single-molecule force-spectroscopy data presented here show: 1) how a redox environment can modulate the mechanical properties of angiostatin; 2) how this modulation relies, at the single-molecule level, on the extent of reduction of the disulfide bonds; and 3) how, at the level of a large sample of molecules, the distribution of the different thiol/ disulfide intermediates after reduction can be estimated by statistical analysis of the force curves.

Optical sensors based on hybrid aptamer/conjugated polymer complexes

Single-stranded DNA (ss-DNA) can specifically bind to various targets, including a complementary ss-DNA, ions, proteins, drugs, and so forth. When binding takes place, the oligonucleotide probe often undergoes a conformational transition. This conformational change of the negatively charged ss-DNA can be detected by using a water-soluble, cationic polythiophene derivative, which transduces the complex formation into an optical (colorimetric or fluorometric) signal without any labeling of the probe or the target. This simple and rapid methodology has enabled the specific and sensitive detection of nucleic acids and human thrombin. This new biophotonic tool can easily be applied to the detection of various other biomolecules and is also useful in the high-throughput screening of new drugs.

Detection of target DNA using fluorescent cationic polymer and peptide nucleic acid probes on solid support

BackgroundNucleic acids detection using microarrays requires labelling of target nucleic acids with fluorophores or other reporter molecules prior to hybridization.ResultsUsing surface-bound peptide nucleic acids (PNA) probes and soluble fluorescent cationic polythiophenes, we show a simple and sensitive electrostatic approach to detect and identify unlabelled target nucleic acid on microarray.ConclusionThis simple methodology opens exciting possibilities for applied genetic analysis for the diagnosis of infections, identification of genetic mutations, and forensic inquiries. This electrostatic strategy could also be used with other nucleic acid detection methods such as electrochemistry, silver staining, metallization, quantum dots, or electrochemical dyes.