Polypyrrole based DNA hybridization assays: study of label free detection processes versus fluorescence on microchips

Livache, Thierry; Maillart, Emmanuel; Lassalle, N.; Mailley, Pascal; Corso, Bruno; Guedon, Philippe; Roget, André; Lévy, Yves

doi:10.1016/s0731-7085(03)00176-6

Cited by 62 publications

(39 citation statements)

References 25 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Peaks characteristic of the maleimide (C 5 Figure 4a) and the maleimide ring to which a sulfur had been added (m/z = 127.98, Figure 4b) in the negative ion spectra. After DNA immobilization, the normalized intensity for the reacted maleimide species (C 4 H 2 NO 2 S − , m/z = 127.98) increased and normalized intensity for the unreacted maleimide species (C 4 H 2 NO 2 − , m/z = 96.01) decreased.…”

Section: Tof-sims Analysismentioning

confidence: 99%

Structure and DNA Hybridization Properties of Mixed Nucleic Acid/Maleimide−Ethylene Glycol Monolayers

et al. 2007

View full text Add to dashboard Cite

The surface structure and DNA hybridization performance of thiolated single-strand DNA (HSssDNA) covalently attached to a maleimide-ethylene glycol disulfide (MEG) monolayer on gold have been investigated. Monolayer immobilization chemistry and surface coverage of reactive ssDNA probes were studied by X-ray photoelectron spectroscopy (XPS) and time-of-flight secondary ion mass spectrometry (ToF-SIMS). Orientation of the ssDNA probes was determined by near edge X-ray absorption fine structure (NEXAFS). Target DNA hybridization on the DNA-MEG probe surfaces was measured by surface plasmon resonance (SPR) to demonstrate the utility of these probe surfaces for detection of DNA targets from both purified target DNA samples and complex biological mixtures such as blood serum. Data from complementary techniques showed that immobilized ssDNA density is strongly dependent on the spotted bulk DNA concentration and buffer ionic strength. Variation of the immobilized ssDNA density had a profound influence on the DNA probe orientation at the surface and subsequent target hybridization efficiency. With increasing surface probe density, NEXAFS polarization dependence results (followed by monitoring the N 1s → π* transition) indicate that the immobilized ssDNA molecules reorient towards a more upright position on the MEG monolayer. SPR assays of DNA targets from buffer and serum showed that DNA hybridization efficiency increased with decreasing surface probe density. However, target detection in serum was better on the "high density" probe surface than on the "high efficiency" probe surface. The amount of target detected for both ssDNA surfaces were several orders of magnitude poorer in serum than in purified DNA samples due to non-specific serum protein adsorption onto the sensing surface.

show abstract

Section: Tof-sims Analysismentioning

confidence: 99%

Structure and DNA Hybridization Properties of Mixed Nucleic Acid/Maleimide−Ethylene Glycol Monolayers

et al. 2007

View full text Add to dashboard Cite

show abstract

“…33 For comparison, a typical fluorescence-based assay is limited by background to a detection limit of 1 pM for DNA hybridization adsorption. 31,34,35 In addition, at a target concentration of 1 pM the surface signal is limited by both adsorption kinetics (eq 5) and diffusion to the surface.…”

Section: A Langmuir Adsorption and Desorption Kineticsmentioning

confidence: 99%

Creating Advanced Multifunctional Biosensors with Surface Enzymatic Transformations

2006

View full text Add to dashboard Cite

This article summarizes our recent work on the coupling of surface enzyme chemistry and bioaffinity interactions on biopolymer microarrays for the creation of multiplexed biosensors with enhanced selectivity and sensitivity. The surface sensitive techniques of surface plasmon resonance imaging (SPRI) and surface plasmon fluorescence spectroscopy (SPFS) are used to detect the surface enzymatic transformations in real time. Three specific examples of novel coupled surface bioaffinity/ surface enzymatic processes are demonstrated: (i) a surface enzymatic amplification method utilizing the enzyme ribonuclease H (RNase H) in conjunction with RNA microarrays that permits the ultrasensitive direct detection of genomic DNA at a concentration of 1 fM without labeling or PCR amplification, (ii) the use of RNA-DNA ligation chemistry to create renewable RNA microarrays from single stranded DNA microarrays, and (iii) the application of T7 RNA polymerase for the onchip replication of RNA from double stranded DNA microarray elements. In addition, a simple yet powerful theoretical framework that includes the contributions of both enzyme adsorption and surface enzyme kinetics is used to quantitate surface enzyme reactivity. This model is successfully applied to SPRI and SPFS measurements of surface hydrolysis reactions of RNase H and Exonuclease III (Exo III) on oligonucleotide microarrays.

show abstract

“…Our previous work focused on the creation and characterization of robust surface chemistries for the attachment of various biomolecules onto chemically modified gold surfaces with a novel methodology based on polypyrrole-based surface functionalization (Guedon et al, 2000). We also showed that multispot parallel DNA hybridization and denaturation could also be detected (Livache et al, 2003), unlike the commercially available instruments that are limited to four channel detection.…”

Section: Published Online 7 June 2004mentioning

confidence: 99%

Versatile analysis of multiple macromolecular interactions by SPR imaging: application to p53 and DNA interaction

et al. 2004

Self Cite

View full text Add to dashboard Cite

The greatest challenge in the postgenomic era is the description of proteome interactions, such as protein-protein or protein-DNA interactions. Surface plasmon resonance (SPR) is an optical technique in which binding of an analyte to the surface changes the refractive index at the surface/solution interface. Molecular interactions are analysed in real time without a labeling step. Currently, the limit to SPR imaging is the small number of reactions that can be simultaneously analysed. Using a novel grafting technology and a new imaging system, we increased the throughput of SPR imaging. The interaction between p53 and DNA was chosen as a paradigm for validation of this assay. Using a tagged DNA methodology, we simultaneously targeted multiple DNA sequences on a single chip. The interaction between p53 and these DNA sequences was monitored by SPR imaging. Qualitative and quantitative analysis provides results similar to those obtained with conventional technologies.

show abstract

Polypyrrole based DNA hybridization assays: study of label free detection processes versus fluorescence on microchips

Cited by 62 publications

References 25 publications

Structure and DNA Hybridization Properties of Mixed Nucleic Acid/Maleimide−Ethylene Glycol Monolayers

Structure and DNA Hybridization Properties of Mixed Nucleic Acid/Maleimide−Ethylene Glycol Monolayers

Creating Advanced Multifunctional Biosensors with Surface Enzymatic Transformations

Versatile analysis of multiple macromolecular interactions by SPR imaging: application to p53 and DNA interaction

Contact Info

Product

Resources

About