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. 2016 Jul 7;6:28711. doi: 10.1038/srep28711

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Copyright © 2016, Macmillan Publishers Limited

This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/

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Samples were incubated with PK and subjected in different dilutions (1:1000 to 1:1000000) to a membrane adsorption assay. PK-resistant PrP^res was detected via 3F4 staining. (A) Lanes 1–4 show the filtation of RT-QuIC reaction mix seeded with brain homogenate from sCJD patients (MM1) (diluted 10⁻³) before and after the RT-QuIC run. Treatment with doxycycline (1 mM) reduced the amount of PK resistant PrP after RT-QuIC significantly (lane 4) compared to the untreated reaction (lane 3). (B) Brain homogenates from sCJD MM1 patients were incubated with different doxycycline concentrations (0–10 mM) and analysed via membrane absorption assay. Lanes 1–3 indicate that the amount of PK-resistant PrP remained unchanged after doxycycline treatment. No PK-resistant PrP was detected in the negative control consisting of brain homogenate (10%) from patients without prion disease (lane 4). Brain homogenates (10%) from sCJD MM1 patients revealed PK resistant PrP (lane 5). For confirmation we repeated the experiment three times with n = 3 patients.