Table 2.
Extraction Buffer | Sample: Buffer Ratio | Additional Components Added | Incubation Time | Viral Titer Post-Treatment (log CCID50/ 0.1 mL) |
---|---|---|---|---|
PBS (control) | 1:4 | None | 30 min | 8.6 ± 0.1 |
Trizol LS | 1:3 | None | 30 min | Below LOD |
Viral RNA Buffer | 1:3 | 0.5 % β-mercaptoethanol | 30 min | Below LOD |
Buffer AVL | 1:4 | None | 30 min | 3.6 ± 0.4 |
Buffer AVL + Ethanol | 1:4 | Ethanol (44 % final) | 30 min | Below LOD |
UNEX | 1:1 | None | 30 min | 1.4 ± 0.1 |
UNEX + Ethanol | 1:1 | Ethanol (50 % final) | 30 min | Below LOD |
MagMAX Lysis/Binding Concentrate | 1:2 | None | 30 min | Below LOD |
High Pure Binding Buffer | 1:2 | None | 30 min | 7.8 ± 0.3 |
High Pure Binding Buffer with Proteinase K | 1:2 | Proteinase K | 30 min | Below LOD |
WPV1 isolate was incubated with extraction buffers Buffer AVL, Trizol and UNEX, Viral RNA Buffer, MagMAX Lysis/Binding Concentrate, or High Pure Binding Buffer with or without ethanol and reducing agents, for 30 min at room temperature. Samples were concentrated and resulting viral titers determined by CCID50 assay. LOD, Limit of detection (0.5 log10 CCID50/0.1 mL).