Related Experiment Video
Updated: Aug 15, 2026
Real-time Quaking-induced Conversion Assay for Detection of CWD Prions in Fecal Material
Published on: September 29, 2017
An immunoassay for the pathological form of the prion protein based on denaturation and time resolved fluorometry
Reza H Dabaghian1, Geoff Barnard, Ian McConnell
1Virus Reference Department, Centre for Infections, Health Protection Agency, 61 Colindale Avenue, London NW9 5HT, UK. Reza.Dabaghian@hpa.org.uk
Abstract:
Concern about the possible secondary spread of variant Creutzfeldt-Jakob disease (vCJD) through blood transfusion and blood products has increased the need for a sensitive and rapid test for the identification of PrP(Sc) in specimens collected non-invasively from living persons. Furthermore, an accurate estimate of the prevalence of pre-clinical vCJD in the British population would be possible if there were such a test that could be applied to specimens available readily (e.g. blood and urine). As a first step towards that goal, we have developed a simple and sensitive test for the detection of PrP(Sc) in peripheral tissues and brain of vCJD patients, based on the differential extraction of PrP(Sc) with guanidine hydrochloride. The prion protein (PrP) isoforms are extracted sequentially from homogenized tissue by applying two different concentrations of this chaotropic agent. Each extraction yields a fraction of the PrP isoforms with different solubilities in guanidine hydrochloride. Quantitation of the two fractions (relatively insoluble or relatively soluble) using time resolved fluorescence (DELFIA) as a reporter system allows differentiation between PrP(Sc) infected and non-infected tissues. The assay has a detection limit of 10 pg PrP, is robust and could be automated.
Insights
A new test detects PrP(Sc), the infectious agent of variant Creutzfeldt-Jakob disease (vCJD), in peripheral tissues. This sensitive assay uses differential guanidine hydrochloride extraction and fluorescence detection, paving the way for non-invasive vCJD diagnostics.
Area of Science:
- Neuroscience
- Biochemistry
- Medical Diagnostics
Background:
- Variant Creutzfeldt-Jakob disease (vCJD) poses a risk for secondary transmission via blood products.
- A sensitive, rapid diagnostic test for vCJD in living individuals is crucial for public health.
- Current diagnostic methods are limited, especially for pre-clinical detection in non-invasive samples.
Purpose of the Study:
- To develop a sensitive and rapid test for detecting PrP(Sc) in vCJD patients.
- To establish a method for identifying PrP(Sc) in non-invasively collected specimens.
- To enable accurate prevalence estimation of pre-clinical vCJD.
Main Methods:
- Developed a test based on differential extraction of PrP(Sc) using guanidine hydrochloride.
- Sequentially extracted PrP isoforms from homogenized tissues at two guanidine hydrochloride concentrations.
- Quantified PrP fractions using time-resolved fluorescence (DELFIA) for differentiation.
Main Results:
- The assay successfully detects PrP(Sc) in peripheral tissues and brain samples from vCJD patients.
- The method demonstrates a detection limit of 10 pg PrP.
- The assay is robust and has potential for automation.
Conclusions:
- A simple, sensitive test for PrP(Sc) detection in vCJD has been developed.
- This assay is a significant first step towards non-invasive vCJD diagnostics.
- The test's sensitivity and potential for automation offer promising avenues for vCJD screening and prevalence studies.
