The pattern of monoclonal antibody recognition of IEs varied between parasite isolates. This is consistent with the finding that the var2csa sequence is composed of conserved stretches separated by stretches with substantial interclonal diversity (Duffy et al., 2006a; Trimnell et al., 2006) , and the prediction that B-cell epitopes in VAR2CSA DBL3-X locate mainly to polymorphic, surface-exposed parts of VAR2CSA . We therefore cloned and sequenced 43 VAR2CSA DBL3-X domains from placental parasite isolates. A subset of 29 of these domains selected to represent the overall VAR2CSA DBL3-X diversity was expressed as Baculovirus recombinant proteins and used in ELISA to test the specificity of the three VAR2CSA DBL3-X-specific monoclonals. The PAM2.8 antibody reacted with 25, PAM6.1 with eight and PAM8.1 with 20 of the domain variants (Fig. 4A) . A multiple sequence alignment of all the proteins indicated that the main difference between the PAM8.1-negative and -positive proteins was a C-terminal 16-amino-acid stretch that maps to a polymorphic region of 3D7-VAR2CSA DBL3-X, which is predicted to be a surfaceexposed loop (Fig. 4B) . Residues in this region either were deleted in the PAM8.1-negative proteins or had a different amino acid composition compared with the PAM8.1-positive variants (Fig. 4A) . To substantiate this possibility we constructed a chimeric protein where the 16-amino-acid stretch from a PAM8.1-positive domain variant (FCR3) was transferred to the corresponding site in a PAM8.1-negative variant lacking this sequence (3D7) (Fig. 4A, bottom) . The recombinant proteins corresponding to the unmodified FCR3 sequence and the chimeric construct both tested positive in Western blots probed with PAM8.1, in contrast to the recombinant protein representing the authentic 3D7 sequence (Fig. 4C) , thus confirming the predicted position of the PAM8.1 epitope. It was not possible to predict the exact targets of PAM2.8 and PAM6.1 by multiple alignments of the primary sequences.