Abstract
Identification of P. falciparum infected erythrocyte surface ligands (such as PfEMP1) matched with the host receptors they interact with, as well as identification of PfEMP1 domains that are targets of protective immunity, are important for understanding of the pathophysiology of severe malaria (SM) and for design of novel vaccine candidates. In addition, identification of small-molecule drugs that can prevent or reverse receptor-ligand domain interactions could provide new tools for adjunctive therapy in SM. This protocol describes how to prepare functionally intact PfEMP1 proteins in mammalian cells (COS-7) and immobilize them on the surface of BioPlex beads. Furthermore, the protocol described how to identify PfEMP1 constructs that bind to specific host receptors or to immunoglobulins (IgG, IgM, etc.), and how to measure inhibition of the receptor binding to PfEMP1 constructs by small-molecule compounds or serum/plasma.
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Acknowledgments
I am truly thankful to a number of people, who contributed hands-on to the development, testing, and using these protocols, these are Eddy Rossnagle, Emily Amos, Tyler I. Frey, Valentina Voronkova, Tracy Saveria, Justin Gullingsrud, Olga Chesnokov, and Sergey O. Tcherniuk. I especially grateful to Emily Amos and Justin Gullingsrud who prepared a number of detailed notes for the lab protocols that I have used in preparation of this chapter. I am also thankful to Jonathan Kurtis, who convinced me long time ago to switch to BioPlex protein array system from a planar format, and to Patrick Duffy for his constant support (intellectual and financial) of the development of this functional protein array system for studies of surface antigens of malarial parasites.
Funding was provided by the Bill & Melinda Gates Foundation (grants 29202 and 1634); NIH (grants 1R21AI064503, 1R56AI083668, 1R01AI092120, 1R21AI137721), and Florida Atlantic University start-up fund.
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Oleinikov, A.V. (2022). High-Throughput BioPlex Assay for the Study of Functionally Active Plasmodium Falciparum Antigens That Are Expressed on the Surface of Infected Erythrocytes. In: Jensen, A.T.R., Hviid, L. (eds) Malaria Immunology. Methods in Molecular Biology, vol 2470. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2189-9_24
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DOI: https://doi.org/10.1007/978-1-0716-2189-9_24
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