The PMCA assay was optimized for adaptation to low level detection of PrPSc in hamster plasma. Evaluation of numerous key variables of the PMCA assay led to an optimized protocol capable of ~3 log10 amplification after 32 cycles (two 16 hour rounds). When commercially purchased normal hamster plasma was added to the PMCA reaction an accentuation in PrPSc amplification was observed (>6.75 log10 after 32 cycles). Only con-specific plasma appeared to enhance the conversion of PrPC to PrPSc, suggesting that a species-specific co-factor may be involved in assembly of protein aggregates. Serial PMCA in the presence of low level (10%) contiguous conspecific plasma resulted in the generation of de novo PrPSc after several rounds of PMCA. Although plasma significantly accentuated PrPSc amplification by PMCA, the formation of de novo PrPSc interfered with the ability of using the PMCA assay to detect prion infections in hamsters experimentally infected with 263K scrapie. / Physiology, Cell and Developmental Biology
| Identifer | oai:union.ndltd.org:LACETR/oai:collectionscanada.gc.ca:AEU.10048/1407 |
| Date | 11 1900 |
| Creators | Braithwaite, Shannon Lynn |
| Contributors | Belosevic, Miodrag (Biological Sciences), Neumann, Norman (School of Public Health), Allison, Ted (Biological Sciences), Aiken, Judd (Agriculture, Food & Nutritional Sciences), McKenzie, Debbie (Biological Sciences) |
| Source Sets | Library and Archives Canada ETDs Repository / Centre d'archives des thèses électroniques de Bibliothèque et Archives Canada |
| Language | English |
| Detected Language | English |
| Type | Thesis |
| Format | 1388574 bytes, application/pdf |
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