Reactivities of antibodies to P-proteins.

<p>(<b>A</b>) Schematic of the full length <i>P. falciparum</i> P2 protein and different deletion constructs (PfP2CΔ20, PfP2CΔ40). The epitope positions of P2, where the monoclonals E2G12 and A12D9 map, are shown. (<b>B</b>) <b>I:</b> ELISA using 100 ng of each recombinant PfP2, PfP2CΔ40 and PfP1 proteins, against a panel of 7 anti-PfP2 monoclonal antibody supernatants (neat) and a rabbit α-PfP1-polyclonal antibody (1∶ 2000 dilution). <b>II:</b> Titration of polyclonal anti-PfP1 antisera against 100 ng PfP1 and PfP2 proteins. <b>III:</b> ELISA of 7 different PfP2 peptides (details described in <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1002858#s4" target="_blank">Methods</a> section) to map the epitope of E2G12. It binds to peptide 5, amino acid 27 to 49 (H-NVLGA VNADV EDEVL NNFID SLK-OH). (<b>C</b>) About 4 µg each of recombinant proteins-rPfP2, rPfP2CΔ20, rPfP2CΔ40, GST-PfP1, GST-PfP0C and GST-were resolved on 12% SDS-PAGE. (a) Coomassie stain of the gel; (b), (c) and (d) are immunoblots probed with (b) α-P0 mAb E5F4 <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1002858#ppat.1002858-Uchiumi1" target="_blank">[36]</a>; (c) α-P2 mAb A12D9; and (d) α-P2 mAb E2G12. (<b>D</b>) Immunoblots of 80 µg of <i>P. falciparum</i>, <i>P. berghei</i> and <i>P. yoelii</i> total parasite lysates probed with (I) α-P2 mAb E2G12 and α-P0 mAb E5F4; (II) Immunoblots of gradient SDS-PAGE of <i>P. falciparum</i> parasite crude protein, probed with α-P2 mAb E2G12 and α-PfP1-polyclonal antibodies; (III) Immunoblots of 50 µg each of <i>P. falciparum</i>, <i>Toxoplasma gondii</i>, mouse and human protein extracts, probed with E2G12. Actin was used as a loading control.