SBIR-STTR Award

Adlyfe Inc. has developed a highly sensitive fluorescence based test, called the Misfolded Protein Diagnostic (MPD) Assay, for the misfolded prion protein associated with infectivity. The femtomolar sensitivity of the test enables the presymptomatic detection in blood, which can be performed ante mortem in routine surveillance of live animals and in blood screening of the human blood supply. The MPD Assay does not rely on antibodies but mimics the folding activity of prions utilizing a target binding peptide. The amplification of the signal through nucleation of folding of other added binding peptides, allows for the extreme sensitivity of the detection method to be realized. This has been demonstrated in controlled in vitro biophysical studies, controlled models of disease in hamsters and mice, and in endemic disease with blood samples from scrapie sheep, BSE-infected cows and monkey and human sCJD. Current protocols and methodology have been geared toward laboratory bench execution of the MPD assay. Here, we propose to move this exciting new technology forward to a more robust integrated assay system that will ultimately encompass software for data analysis and reporting.

Keywords:
Prion Diagnostics, Peptide-Based Diagnostics, Misfolded Protein, Conformational Change

The prion disease process involves a conformational change in the prion protein that in turn can serve as the basis for an early detection, diagnostic, prognostic and screening assay. The Misfolded Protein Diagnostic (MPD) Assay detects infectivity in a crude sample through the use of labeled Pronucleon peptides that undergo conformational change in the presence of the misfolded infectious prion protein. The conformational peptide change is propagated throughout the ensemble of sequence specific Pronucleon peptides in the reaction via a nucleation event resulting in an amplification of signal. Preliminary data supports the simplicity and exquisite sensitivity of this non-immunological approach to infectious prion protein detection. Level of detection has been determined to be near 1 infectious dose for titered murine sCJD plasma samples. During Phase I funding, the MPD Assay has been under optimization with keen focus on defining the positive controls for the assay as well as looking at variability between normal human plasma samples to better define the threshold of detection. With continued funding we propose to define and further optimize the assay reagents and metrics into a platform system that can be reduced to a high-throughput, small volume automated format.

Keywords:
Prions, Prion Diagnostic, Misfolded Protein, Tse Diagnostic, Prp