Figure 1

The capacity of polymeric and monomeric NP to bind mAb N5D3. A) RIPA. Radiolabeled cytosol of infected cells was divided into unheated (r.t) portion, containing NP polymers and the heated (70° C) portion, containing NP 56 kDa monomers as a result of NP polymers dissociation. Both unheated and pre-heated portions were subjected to RIPA using polyclonal anti-NP Abs or mAb N5D3. SDS-PAGE of the immunoprecipitates obtained by RIPA using polyclonal Abs (lanes 1, 2) and mAb N5D3 (lanes 3,4). B) Immunolotting. Radiolabeled unheated (1,3) and pre-heated (2,4) cytosols were subjected to SDS-PAGE, followed by Western blot, including electro-transfer onto nitrocellulose membrane, autoradiography and immunodetection using mAb N5D3. Autoradiography (lanes 1, 2) and immunostaining using mAb N5D3 (lanes 3,4) of membrane containing blotted proteins. C) Renaturation of N5D3 epitope caused by self-association of the concentrated soluble NP monomers. The non-concentrated m-NP before RIPA (lane 1) and after immunosorption by RIPA using mAb N5D3 (lane 2). The concentrated soluble self-associated m-NP (as described in the text) before RIPA (lane 3) and after immunosorption by RIPA using mAb N5D3 (lane 4). The aliquot of RIPA immunoprecipitate shown in lane 4 was heated at 100°C for 3 min before SDS-PAGE (lane 5). The samples shown in lanes 1–4 were not additionally pre-heated before SDS-PAGE.