title: Development and Application of a Simple Plaque Assay for the Human Malaria Parasite Plasmodium falciparum.
abstract: Malaria is caused by an obligate intracellular protozoan parasite that replicates within and destroys erythrocytes. Asexual blood stages of the causative agent of the most virulent form of human malaria, Plasmodium falciparum, can be cultivated indefinitely in vitro in human erythrocytes, facilitating experimental analysis of parasite cell biology, biochemistry and genetics. However, efforts to improve understanding of the basic biology of this important pathogen and to develop urgently required new antimalarial drugs and vaccines, suffer from a paucity of basic research tools. This includes a simple means of quantifying the effects of drugs, antibodies and gene modifications on parasite fitness and replication rates. Here we describe the development and validation of an extremely simple, robust plaque assay that can be used to visualise parasite replication and resulting host erythrocyte destruction at the level of clonal parasite populations. We demonstrate applications of the plaque assay by using it for the phenotypic characterisation of two P. falciparum conditional mutants displaying reduced fitness in vitro.
authors.affiliation: The Francis Crick Institute, Mill Hill Laboratory, Mill Hill, London, NW7 1AA, United Kingdom.
authors.forename: James A
authors.initials: JA
authors.lastname: Thomas
authors.affiliation: The Francis Crick Institute, Mill Hill Laboratory, Mill Hill, London, NW7 1AA, United Kingdom.
authors.forename: Christine R
authors.initials: CR
authors.lastname: Collins
authors.affiliation: The Francis Crick Institute, Mill Hill Laboratory, Mill Hill, London, NW7 1AA, United Kingdom.
authors.forename: Sujaan
authors.initials: S
authors.lastname: Das
authors.affiliation: The Francis Crick Institute, Mill Hill Laboratory, Mill Hill, London, NW7 1AA, United Kingdom.
authors.forename: Fiona
authors.initials: F
authors.lastname: Hackett
authors.affiliation: Department of Microbiology & Molecular Medicine, University of Geneva, Rue Michel-Servet 1, CH-1211, Geneva 4, Switzerland.
authors.forename: Arnault
authors.initials: A
authors.lastname: Graindorge
authors.affiliation: The Francis Crick Institute, Mill Hill Laboratory, Mill Hill, London, NW7 1AA, United Kingdom.
authors.forename: Donald
authors.initials: D
authors.lastname: Bell
authors.affiliation: The Francis Crick Institute, Mill Hill Laboratory, Mill Hill, London, NW7 1AA, United Kingdom.
authors.forename: Edgar
authors.initials: E
authors.lastname: Deu
authors.affiliation: The Francis Crick Institute, Mill Hill Laboratory, Mill Hill, London, NW7 1AA, United Kingdom.|Department of Pathogen Molecular Biology, London School of Hygiene and Tropical Medicine, London, WC1E 7HT, United Kingdom.
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Blackman
mesh_terms: D000818:Animals; D004912:Erythrocytes / Q000469:parasitology; D006462:Hemolytic Plaque Technique / Q000379:methods*; D006801:Humans; D008018:Life Cycle Stages; D016778:Malaria, Falciparum / Q000469:parasitology*; D020066:Merozoite Surface Protein 1 / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D010271:Parasites / Q000302:isolation & purification*; D010641:Phenotype; D010963:Plasmodium falciparum / Q000254:growth & development / Q000302:isolation & purification*
chemical_list: D020066:Merozoite Surface Protein 1