Preparation of Conjugates between Oligonucleotides and <i>N</i>-Vinylpyrrolidone/<i>N</i>-Acryloxysuccinimide Copolymers and Applications in Nucleic Acid Assays To Improve Sensitivity

Erout, Marie-Noëlle; Troesch, Alain; Pichot, Christian; Cros, Philippe

doi:10.1021/bc960046a

Cited by 47 publications

(53 citation statements)

References 25 publications

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“…The tests were performed on bioMérieux's Vidas immunoanalyzer, but the protocol was greatly modified from the one reported in [3,10]. In our procedure, an intermediate detection probe (see Fig.…”

Section: Hbv Virus Dna Detectionmentioning

confidence: 99%

“…The other part of the intermediate, which was not complementary to the HBV genome, was used for hybridization with the detection polymer. The advantage of this protocol, as compared with the one described in [10], lies in the fact that it allows a so-called 'universal' detection conjugate to be used, i.e. one that can be applied to many other tests, irrespective of the genome to be detected.…”

Section: Hbv Virus Dna Detectionmentioning

confidence: 99%

“…In this context, in order to meet the high sensitivity requirements, signal amplification has to be achieved, and our approach has relied on the use of polymer/ODN conjugates. In this section, the data on the comparative efficiency of conjugates obtained with the saccharidic homo and copolymers with those obtained with a (N-vinylpyrrolidone-alt-N-acryloxysuccinimide) copolymer (NVP/NAS) [10] are reported.…”

Section: Application To the Detection Of The Dna Of Hepatitis B Virusmentioning

confidence: 99%

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Conjugation of nucleic acid probes to 6-aminoglucose-based homo- and copolymers. II. Application to diagnostics

Delair

Badey

Charles

et al. 1997

Polym. Adv. Technol.

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The covalent immobilization of oligodeoxynucleotides to a 6‐deoxy‐6‐methacryloylamido‐d‐glucopyranose (6‐amino glucose) homopolymer and a 36% hydroxyethylmethacrylate (HEMA) copolymer was studied in order to make oligodeoxynucleotide–polymer conjugates of potential applications in diagnostics. The reducing ends of the saccharidic moieties of the (co)polymers served as reactive groups for the covalent attachment of 5′‐amino‐modified oligodeoxyribonucleotides via reductive amination. The course of the immobilization reaction was slightly affected by the polymer composition, but differences were marked as far as the aggregation of the conjugates was concerned. Evaluation of the conjugates in the quantification of hepatitis B virus DNA allowed amplification factors identical to the reference polymer. Amplification was the most efficient with conjugates obtained, at low pH, with the copolymer. © 1997 John Wiley & Sons, Ltd.

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Section: Hbv Virus Dna Detectionmentioning

confidence: 99%

Section: Hbv Virus Dna Detectionmentioning

confidence: 99%

Section: Application To the Detection Of The Dna Of Hepatitis B Virusmentioning

confidence: 99%

See 1 more Smart Citation

Conjugation of nucleic acid probes to 6-aminoglucose-based homo- and copolymers. II. Application to diagnostics

Delair

Badey

Charles

et al. 1997

Polym. Adv. Technol.

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“…Also, conjugates based on poly(N-vinyl pyrrolidone-co-N-acryloxysuccinimide) [poly(NVP/NAS)] enhanced sensitivity when used at the detection phase. 9 Indeed, they multiplied the number of detection probes, inducing a higher colorimetric or fluorescent signal. In both cases, the conjugates were prepared via the covalent coupling of amino-bearing ODNs (independently presynthesized) onto a reactive polymer, which often resulted in aggregate formation [compact random coil; number-average molecular weight (M n ) Ϸ 2,000,000 g/mol].…”

Section: Introductionmentioning

confidence: 99%

Oligonucleotide synthesis onto poly(N‐acryloylmorpholine‐co‐N‐acryloxysuccinimide): Assessment of the resulting conjugates in a DNA sandwich hybridization test

Minard‐Basquin

Chaix

D’Agosto

et al. 2004

J of Applied Polymer Sci

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ABSTRACT:A water-soluble statistical poly(N-acryloylmorpholine-co-N-acryloxysuccinimide) [poly(NAM/NAS)] copolymer was studied for polymer-oligonucleotide (ODN) conjugate elaboration and for further use in diagnostic applications. Three different copolymers were first prepared by free-radical solution polymerization with different Nacryloylmorpholine (NAM) and N-acryloxysuccinimide (NAS) molar ratios (80/20, 70/30, and 60/40). Their number-average molecular weights ranged from 98,000 to 120,000 g/mol, as determined by aqueous size exclusion chromatography with an online light-scattering detector. Then, polymer-ODN conjugates were obtained via a strategy consisting of the direct synthesis of ODNs onto polymer chains previously grafted onto a controlled pore glass support. Before the grafting of the polymer onto the solid support, a preliminary step was performed to bind a nucleotide starter along the polymer chain (via the reactive NAS units) to initiate automated DNA synthesis. To multiply the number of ODNs growing from starters, a branched phosphoramidite synthon [bearing two O-dimethoxytrityl groups] was introduced at the first step of ODN elongation as a short sequence of four branched synthons alternated with three thymidine residues. Conjugates were assessed in a DNA sandwich hybridization test developed for hepatitis B virus detection. Sensitivity limits were evaluated and compared to those obtained with an other polymer, poly(maleic anhydride-alt-methyl vinyl ether) [poly(MA/MVE)]. A sensitivity limit of 2.6 ϫ 10 7 DNA copies/mL was reached with the poly(MA/MVE)-ODN conjugate at the capture phase and with the poly(NAM/NAS)-branched ODN conjugate at the detection phase of the test.

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“…Both of these concepts were described in patents originating from this lab 3,4 and were successfully used for the detection of the hepatitis B virus DNA. 5 Various polymers have been tested, and all of them were based on the use of a reactive comonomer capable of binding amino counterparts by the formation of amide or imine bonds. Hence, linear synthetic functional N-vinyl pyrolidone-N-acryloxy succinimide, 4 -7 maleic anhydride copolymers, 8,9 and glucose amine polymers 10 have been described to covalently bind prepurified oligonucleotides.…”

Section: Introductionmentioning

confidence: 99%

Oligonucleotide synthesis on maleic anhydride copolymers covalently bound to silica spherical support and characterization of the obtained conjugates

Chaix

Minard‐Basquin

Delair

et al. 1998

J. Appl. Polym. Sci.

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ABSTRACT:A new route was proposed to make polymer-oligonucleotide conjugates of potential applications in diagnostics. It consisted in direct synthesis of oligonucleotides onto controlled pore glass surface grafted with poly(maleic anhydride-alt-methyl vinyl ether) (P[MAMVE]) or poly(maleic anhydride-alt-ethylene) (P [MAE]). The anhydride moieties were used for both the covalent coupling of the copolymer via ester bond and binding of 5Ј-dimethoxytrityl thymidine 3Ј-(6-aminohexyl phosphate) (I) initiator of oligodeoxynucleotide (ODN) synthesis via amide bond. The difference of stability between ester and amide links under basic treatment was used for the selective cleavage of (polymer-oligonucleotide) conjugates after DNA synthesis completion. We succeeded in grafting functionalized copolymer onto silica surface and Polythymidine 26-mer ODN was performed. After concentrated ammonium hydroxide treatment, conjugate crude materials were characterized by size exclusion chromatography coupled to multiangle laser light scattering detection. The number average molecular weight (Mn) for conjugate with P[MAMVE] was abnormally lower than expected and was assigned to polymer degradation using high pH conditions. Such a phenomenon did not occur with P[MAE]-polythymidine conjugate. However, in both cases, parasite ODN synthesis was also evidenced, which was attributed to thymidine phosphoramidite adsorption side reaction during DNA synthesis.

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Preparation of Conjugates between Oligonucleotides and N-Vinylpyrrolidone/N-Acryloxysuccinimide Copolymers and Applications in Nucleic Acid Assays To Improve Sensitivity

Cited by 47 publications

References 25 publications

Conjugation of nucleic acid probes to 6-aminoglucose-based homo- and copolymers. II. Application to diagnostics

Conjugation of nucleic acid probes to 6-aminoglucose-based homo- and copolymers. II. Application to diagnostics

Oligonucleotide synthesis onto poly(N‐acryloylmorpholine‐co‐N‐acryloxysuccinimide): Assessment of the resulting conjugates in a DNA sandwich hybridization test

Oligonucleotide synthesis on maleic anhydride copolymers covalently bound to silica spherical support and characterization of the obtained conjugates

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