A sensitive and quantitative assay for normal PrP in plasma.
Transmissible spongiform encephalopathies can be transmitted by blood transfusion. The risk of spreading the disease among the human population could be mitigated with the implementation of a blood screening assay. We developed a two-antibody assay for PrP detection in plasma using the ORIGEN technology with a protocol modification to improve the limit of detection and to increase the sample volume assayed. In the standard 200 microL format, the assay had a detection limit of 7-10 pg of recombinant PrP and 3 pg in 1 mL final volume implementation. PrP concentration measured in normal and scrapie-infected hamster brains was 7.5+/-0.9 and 57.3+/-9.6 microg/g, respectively. After a concentration step with an immuno-affinity resin, plasma PrP(c) was detected by Western blot and its concentration was measured at 3.5+/-0.8 ng/mL. From these data and assuming that blood has the same specific infectivity as brain, we estimated the concentration of abnormal PrP in hamster-infected plasma to be 32 f g/mL. The assay also detected abnormal brain PrP spiked into plasma although the limit of detection was affected. This is a novel and sensitive assay for the detection of PrP in plasma that could be developed into a platform for a plasma-based TSE test.
Gregori
Luisa
L
Veterans Affairs Medical Center and University of Maryland, Baltimore, MD 21201, United States. lgreg002@umaryland.edu
Gray
Benjamin N
BN
Rose
Elaine
E
Spinner
Daryl S
DS
Kascsak
Richard J
RJ
Rohwer
Robert G
RG
D000818:Animals; D015153:Blotting, Western; D006224:Cricetinae; D018562:Disease Transmission, Infectious / Q000517:prevention & control; D006801:Humans; D007118:Immunoassay / Q000379:methods*; D010949:Plasma / Q000737:chemistry*; D017096:Prion Diseases / Q000517:prevention & control; D011328:Prions / Q000097:blood*; D012680:Sensitivity and Specificity
D011328:Prions