Figure 1. Polyclonal Drosophila S2 stable cell lines expressing PfRH5 protein variants.

(A) Schematic of PfRH5 protein variants. All encoded from the N-terminus: a BiP insect signal peptide (green) followed by the ectodomain of PfRH5 (αα 26–526) (blue) followed by a C-terminal His6 tag (red). Protein v1.0 encoded the 7G8 strain sequence which differs from the 3D7 clone sequence by a single polymorphism Y203C (yellow). The other cysteine residues in PfRH5 are indicated by small black boxes (C224, C317, C329, C345 and C351). Amino acid substitutions to remove N-linked glycan sequons are indicated by red asterisks for threonine (T) to alanine (A), and purple asterisks for asparagine (N) to glutamine (Q). The predicted molecular weight (Mw) for each protein is also shown (based on the primary sequence only and not possible N-linked glycans). (B) Following transfection, stable polyclonal Drosophila S2 cell lines were generated encoding PfRH5 protein variants versions 1.0–5.0. 30 μL samples of culture supernatants from batch cultures were run on SDS-PAGE gels under reducing conditions and stained for total protein (right hand side) or Western blots performed with anti-Penta-His mAb (left hand side). Results are shown for each variant. M = molecular weight markers. −ve = S2 cell supernatant control.