Sensitive, preclinical detection of prions in brain by nanospray liquid chromatography/tandem mass spectrometry.
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 have quantitated the amount of proteinase K-resistant prion protein (PrP 27-30) by use of nano-scale liquid chromatography coupled to tandem mass spectrometry 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 h after inoculation. Our results show sensitive detection of 180 fmol of PrP 27-30 per g brain (wet weight) as early as 24 h after inoculation. Clinical symptoms are not observed until 9 weeks after inoculation.
Onisko
Bruce C
BC
Western Regional Research Center, Agricultural Research Service, United States Department of Agriculture, Albany, CA 94710, USA. bonisko@pw.usda.gov
Silva
Christopher J
CJ
Dynin
Irina
I
Erickson
Melissa
M
Vensel
William H
WH
Hnasko
Robert
R
Requena
Jesús R
JR
Carter
John Mark
JM
D000818:Animals; D001921:Brain / Q000821:virology*; D001923:Brain Chemistry; D002851:Chromatography, High Pressure Liquid; D006224:Cricetinae; D004195:Disease Models, Animal; D005260:Female; D008647:Mesocricetus; D036103:Nanotechnology / Q000379:methods*; D018622:PrP 27-30 Protein / Q000032:analysis*; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000379:methods*
D018622:PrP 27-30 Protein