title: A rapid and scalable density gradient purification method for Plasmodium sporozoites.
abstract: BACKGROUND
  Malaria remains a major human health problem, with no licensed vaccine currently available. Malaria infections initiate when infectious Plasmodium sporozoites are transmitted by Anopheline mosquitoes during their blood meal. Investigations of the malaria sporozoite are, therefore, of clear medical importance. However, sporozoites can only be produced in and isolated from mosquitoes, and their isolation results in large amounts of accompanying mosquito debris and contaminating microbes.
  
  
  METHODS
  Here is described a discontinuous density gradient purification method for Plasmodium sporozoites that maintains parasite infectivity in vitro and in vivo and greatly reduces mosquito and microbial contaminants.
  
  
  RESULTS
  This method provides clear advantages over previous approaches: it is rapid, requires no serum components, and can be scaled to purify >107 sporozoites with minimal operator involvement. Moreover, it can be effectively applied to both human (Plasmodium falciparum, Plasmodium vivax) and rodent (Plasmodium yoelii) infective species with excellent recovery rates.
  
  
  CONCLUSIONS
  This novel method effectively purifies viable malaria sporozoites by greatly reducing contaminating mosquito debris and microbial burdens associated with parasite isolation. Large-scale preparations of purified sporozoites will allow for enhanced in vitro infections, proteomics, and biochemical characterizations. In conjunction with aseptic mosquito rearing techniques, this purification technique will also support production of live attenuated sporozoites for vaccination.
authors.affiliation: Center for Mosquito Production and Malaria Infection Research, Seattle Biomedical Research Institute, 307 Westlake Avenue North, Suite 500, Seattle, WA 98109, USA.
authors.forename: Mark
authors.initials: M
authors.lastname: Kennedy
authors.forename: Matthew E
authors.initials: ME
authors.lastname: Fishbaugher
authors.forename: Ashley M
authors.initials: AM
authors.lastname: Vaughan
authors.forename: Rapatbhorn
authors.initials: R
authors.lastname: Patrapuvich
authors.forename: Rachasak
authors.initials: R
authors.lastname: Boonhok
authors.forename: Narathatai
authors.initials: N
authors.lastname: Yimamnuaychok
authors.forename: Nastaran
authors.initials: N
authors.lastname: Rezakhani
authors.forename: Peter
authors.initials: P
authors.lastname: Metzger
authors.forename: Marisa
authors.initials: M
authors.lastname: Ponpuak
authors.forename: Jetsumon
authors.initials: J
authors.lastname: Sattabongkot
authors.forename: Stefan H
authors.initials: SH
authors.lastname: Kappe
authors.forename: Jen C C
authors.initials: JC
authors.lastname: Hume
authors.forename: Scott E
authors.initials: SE
authors.lastname: Lindner
mesh_terms: D000818:Animals; D000852:Anopheles / Q000469:parasitology; D002499:Centrifugation, Density Gradient / Q000379:methods*; D004195:Disease Models, Animal; D005260:Female; D006801:Humans; D008099:Liver / Q000469:parasitology; D008288:Malaria / Q000469:parasitology; D051379:Mice; D010274:Parasitology / Q000379:methods*; D010961:Plasmodium / Q000302:isolation & purification* / Q000472:pathogenicity; D010963:Plasmodium falciparum / Q000302:isolation & purification; D010966:Plasmodium vivax / Q000302:isolation & purification; D010967:Plasmodium yoelii / Q000302:isolation & purification; D034101:Sporozoites / Q000166:cytology*; D014774:Virulence