US 2006/0188929 A1Published application

Method capable of being automated for detection of prpres and uses thereof

OwnerNathalie Morel, Christophe Creminon +1inventors

InventorsNathalie Morel, Christophe Creminon, Jacques Grassi

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Abstract

This detection method uses a solid support whereon plasminogen is immobilized, and includes essentially the following steps: (a) preparing the biological sample during which the latter is incubated on a homogenizing buffer which comprises an ionic or nonionic surfactant, a glucose-containing buffer, a saccharose-based buffer and a PBS buffer or in a capture buffer comprising an ionic surfactant; said step optionally comprising incubation with a proteinase K at a final concentration ranging between 1 and 8 μg/ml; (b) capturing the PrPres on the solid support, which is carried out in the presence of a capture buffer, as defined above; (c) a controlled denaturation of the PrPres fixed on the support comprising incubation with a denaturation buffer including at least one chaotropic agent, at a temperature ranging between room temperature and 100° C.; and (d) detecting the denatured PrPres fixed on said support with a PrP protein-specific antibody.

Claims20

Dependent claims

A method for detecting PrPres in a biological sample, using a solid support, in particular microtitration plates or magnetic beads, on which plasminogen is immobilized, which method comprises:

(a) a step of preparing the biological sample. during which step this sample is incubated in a buffer selected from the group consisting of:
(i) buffers for homogenizing the biological sample comprising (1) a buffer selected from the group consisting of buffers comprising at least one surfactant selected from the group consisting of ionic surfactants and nonionic surfactants, a glucose-containing buffer, a sucrose-based buffer and a PBS buffer and (2) optionally, a proteinase K at a final concentration of between 1 and 8 μg/ml, and
(ii) capture buffers comprising at least (1) a surfactant selected from the group consisting of ionic surfactants, and (2) optionally, a proteinase K at a final concentration of between 1 and 8 μg/ml,
(b) a step of capturing PrPres on said solid support, carried out in the presence of a capture buffer as defined above, without PK, by incubation of the biological sample obtained in step (a) with said support on which plasminogen is covalently immobilized;
(c) a step of controlled denaturation of the PrPres attached to said support by means of the plasminogen, comprising incubation of the PrPres with a denaturing buffer comprising at least one chaotropic agent, at a temperature of between ambient temperature and 100° C., and
(d) a step of detecting the denatured PrPres attached to said support, with a PrP protein-specific antibody.

3 more elements

Dependencies are inferred from the claim text.