Immunogenicity of Self-Associated Aggregates and Chemically Cross-Linked Conjugates of the 42 kDa Plasmodium falciparum Merozoite Surface Protein-1

Self-associated protein aggregates or cross-linked protein conjugates are, in general, more immunogenic than oligomeric or monomeric forms. In particular, the immunogenicity in mice of a recombinant malaria transmission blocking vaccine candidate, the ookinete specific Plasmodium falciparum 25 kDa protein (Pfs25), was increased more than 1000-fold when evaluated as a chemical cross-linked protein-protein conjugate as compared to a formulated monomer. Whether alternative approaches using protein complexes improve the immunogenicity of other recombinant malaria vaccine candidates is worth assessing. In this work, the immunogenicity of the recombinant 42 kDa processed form of the P. falciparum merozoite surface protein 1 (MSP142) was evaluated as a self-associated, non-covalent aggregate and as a chemical cross-linked protein-protein conjugate to ExoProtein A, which is a recombinant detoxified form of Pseudomonas aeruginosa exotoxin A. MSP142 conjugates were prepared and characterized biochemically and biophysically to determine their molar mass in solution and stoichiometry, when relevant. The immunogenicity of the MSP142 self-associated aggregates, cross-linked chemical conjugates and monomers were compared in BALB/c mice after adsorption to aluminum hydroxide adjuvant, and in one instance in association with the TLR9 agonist CPG7909 with an aluminum hydroxide formulation. Antibody titers were assessed by ELISA. Unlike observations made for Pfs25, no significant enhancement in MSP142 specific antibody titers was observed for any conjugate as compared to the formulated monomer or dimer, except for the addition of the TLR9 agonist CPG7909. Clearly, enhancing the immunogenicity of a recombinant protein vaccine candidate by the formation of protein complexes must be established on an empirical basis

Acting Oneself as Another: An Actor\u27s Empathy for her Character

What does it mean for an actor to empathize with the character she is playing? We review different theories of empathy and of acting. We then consider the notion of twofoldness (Wollheim), which has been used to characterize the observer or audience perspective on the relation between actor and character (Smith). This same kind of twofoldness or double attunement applies from the perspective of the actor herself who must, at certain points of preparation, distinguish between the character portrayed and her own portrayal effected in her craft. We argue that this concept helps us to understand how the actor can empathize with her character. For the actor who must study and rehearse her character, empathy may begin with higher-order (narrative or imaginative) processes that provide a contextualized understanding of the character. This understanding eventually integrates with more basic empathic processes in her actual performance