Volume 25, Number 1—January 2019
Research
Variable Protease-Sensitive Prionopathy Transmission to Bank Voles
Figure 4
![Immunoblot characteristics of protease-resistant, disease-related prion protein (resPrPD) distribution in phenotypes T1–T3 and controls. Regular and long exposures revealed the overall similarity of the 3-band profiles in T1 and T2, but resPrPD profile, including glycoform representation, differed in the 2 phenotypes with all 3 monoclonal antibodies (Ab) used. T1 included a 7-kDa band, not detected in T2, similar to mobility and Ab immunoreactivity of the T3 7-kDa fragment. The T1 profile matche](/eid/images/18-0807-F4.jpg)
Figure 4. Immunoblot characteristics of protease-resistant, disease-related prion protein (resPrPD) distribution in phenotypes T1–T3 and controls. Regular and long exposures revealed the overall similarity of the 3-band profiles in T1 and T2, but resPrPD profile, including glycoform representation, differed in the 2 phenotypes with all 3 monoclonal antibodies (Ab) used. T1 included a 7-kDa band, not detected in T2, similar to mobility and Ab immunoreactivity of the T3 7-kDa fragment. The T1 profile matched the profile generated in isogenic bank voles inoculated with sCJDMV1 used as human resPrPD type 1 control (ctrl) (lane 2). The T3 profile, visible only after long film exposures, featured a 7-kDa band, but slower migrating bands with variable immunoreactivity were also visible. None of the T1–T3 profiles matched the original VPSPr profile (first lane) although the ≈7-kDa and both 23-kDa and 19-kDa bands were shared with T1 and T2, respectively (compare first with T1 and T2 lanes). The complexity of the native resPrPD profile from VPSPr homogenate is demonstrated by probing with 1E4, a monoclonal Ab to human PrP highly reactive to VPSPr resPrPD (top right panel) (6). Monoclonal Ab 12B2 (middle panels) with high affinity for human resPrPD type 1 confirmed the type 1 characteristics of the resPrPD associated with the T1 phenotype. The small amount of resPrPD type 1 in 1 T2 bank vole probably represents incomplete proteinase K (PK) digestion (lane 5, right panel) (19). Monoclonal Ab SAF84 to the PrP C-terminus, unreactive to human PrP, further underlined the divergence in resPrPD primary structure in T1 and T2 compared with T3. Aside from revealing an additional 13-kDa fragment, strongly detected in T1 and T1 and weakly in T3, SAF84 did not detect the 7-kDa fragment, supporting its internal origin (i.e., cleaved at both N- and C-termini). Uninoculated bank voles were negative for resPrPD. All samples were PK treated. sCJD, sporadic Creutzfeldt-Jakob disease; uninoc., not inoculated; polym., polymorphism; VPSPr, variably protease sensitive prionopathy.
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1These authors contributed equally to this article.