A novel real-time ultrasonic method for prion protein detection using plasminogen as a capture molecule.
BACKGROUND
High resolution ultrasonography (HR-US) can monitor the molecular changes and biochemical interactions between proteins in real-time. The aim of this study was to use HR-US to characterize the real-time interactions between plasminogen coated beads and PrPSc and to determine if this approach could be applied to the identification of animals affected by prion diseases. Plasminogen, immobilized to beads, was used as a capturing tool for PrPSc in brain homogenates from scrapie affected sheep and the binding reaction was monitored in real-time in an ultrasonic cell.


RESULTS
Changes in the ultrasonic parameters suggested that three processes occurred during the incubation: binding, protein-protein network formation and precipitation and that these processes occurred in a concentration dependent manner. Conversely, when homogenates from normal sheep were similarly examined, no evidence for the occurrence of these processes was found indicating the specificity of the interaction between the plasminogen coated beads and PrPSc.


CONCLUSIONS
These results indicate firstly, that the plasminogen coated beads binded selectively to PrPSc and secondly, that a HR-US system can discriminate between scrapie affected and non-affected samples and thus has potential as a tool for the rapid diagnosis for prion diseases. This approach has the significant advantage of not requiring a proteinase K pre-digestion step, which is routinely used in current PrPSc detection assays.
Veterinary Sciences Centre, School of Agriculture, Food Science and Veterinary Medicine, University College Dublin, Belfield, Dublin 4, Repulic of Ireland. carmen.negredo@ucd.ie <carmen.negredo@ucd.ie>
Carmen
C
Negredo
Eoin
E
Monks
Torres
T
Sweeney
D003199:Computer Systems; D007090:Image Interpretation, Computer-Assisted / Q000379:methods*; D010958:Plasminogen / Q000737:chemistry*; D018620:PrPSc Proteins / Q000737:chemistry*; D011485:Protein Binding; D025941:Protein Interaction Mapping / Q000379:methods*; D014463:Ultrasonography / Q000379:methods*
D018620:PrPSc Proteins; D010958:Plasminogen