|Requena, Jesus - UNIVERSITY OF SANTIAGO|
Submitted to: Rapid Communications in Mass Spectrometry
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: October 11, 2007
Publication Date: November 13, 2007
Citation: Onisko, B.C., Silva, C.J., Dynin, I.A., Erickson, M.L., Vensel, W.H., Hnasko, R.M., Requena, J.R., Carter, J.M. 2007. Sensitive, Preclinical Detection of Prions in Brain by nanospray liquid chromatography/tandem mass spectrometry. Rapid Communications in Mass Spectrometry. 21(24):4023-4026. Interpretive Summary: The more sensitive a method is to measure the amount of prions in brain, the earlier the disease can be detected, even before disease symptoms can be seen. This is a serious problem for scrapie in sheep and BSE in cattle. We have developed a new method that is more sensitive than any reported antibody method, and have used this method in a well-studied hamster disease model, to show how sensitive, and how early we could detect disease.
Technical Abstract: More sensitive detection of prions in brain is important because it would allow early detection of disease in young animals and assure a safer food supply. We quantitated the amount of proteinase K-resistant prion protein (PrP 27-30) by use of nano-scale liquid chromatography coupled to a tandem mass spectrometer using the multiple reaction monitoring mode of operation. We used a method based on the detection of VVEQMCTTQYQK (residues 209-220) obtained by reduction, alkylation and digestion with trypsin. Quantitation of the amount of PrP 27-30 in the brains of Syrian hamsters was possible as early as 24 hours after inoculation. Our results show sensitive detection of 180 fmol of PrP 27-30 per g brain (wet weight) as early as 24 hours after inoculation. Clinical symptoms are not observed until 9 weeks after inoculation.