Figure 2

Western blot and ELISA analysis of interaction of mAbs with the PfCP-2.9 mutants. (A) Effects of PfCP-2.9 mutant containing a single amino acid substitution on the binding of the mAbs in Western blot. The mAbs indicated were used to investigate the binding differences between PfCP-2.9 and its mutant with a single amino acid substitution. Same amount of each purified protein was subjected to SDS-PAGE and Western blot. As indicated on the gel, reduced binding of the inhibitory mAb7G was observed in the mutants of M62 (Phe⁴⁹¹ → Ala), M82 (Glu⁵¹¹ → Gln) and M84 (Arg⁵¹³ → Lys) compared to the binding of this mAb to the PfCP-2.9. Similarly, the binding of mAbW9.10 was reduced in M62 and M82, seperately. In addition, the binding of mAb1E1 was reduced in M159 (Gln¹⁴ → Gly) and M188 (Glu⁴³ → Leu), and abolished in M185 (Lys⁴⁰ → Ile). The bindings of two non-inhibitory antibodies, mAb6G and mAbP5-W12, were both reduced in M185 (Lys⁴⁰ → Ile) and abolished in M159 and M188. (B), (C), (D), (E) and (F): Effects of PfCP-2.9 mutant on the binding of the mAbs in ELISA. Proteins coated were standardized by mAb5.2. The mAbs of 1E1, 7G, G11.12, W9.10 and 6G were serially diluted by PBS-M, followed by a standard ELISA. Their binding differences between PfCP-2.9 and its mutated proteins are shown in (B) to (F). Data shown were repeatable in three separate experiments.