Purification of Sarcocystis neurona sporocysts from opossum (Didelphis virginiana) using potassium bromide discontinuous density gradient centrifugation.
This report describes a new, inexpensive procedure for the rapid and efficient purification of Sarcocystis neurona sporocysts from opossum small intestine. S. neurona sporocysts were purified using a discontinuous potassium bromide density gradient. The procedure provides a source of sporocyst wall and sporozoites required for reliable biochemical characterization and for immunological studies directed at characterizing antigens responsible for immunological responses by the host. The examined isolates were identified as S. neurona using random amplified polymorphic DNA primers and restriction endonuclease digestion assays. This method allows the collection of large numbers of highly purified S. neurona sporocysts without loss of sporocyst viability as indicated by propidium iodide permeability and cell culture infectivity assays. In addition, this technique might also be used for sporocyst purification of other Sarcocystis spp.
Elsheikha
Hany M
HM
Molecular Epidemiology Laboratory, Food Safety and Toxicology Center, Michigan State University, East Lansing, MI 48824, USA.
Murphy
Alice J
AJ
Fitzgerald
Scott D
SD
Mansfield
Linda S
LS
Massey
Jeffrey P
JP
Saeed
Mahdi A
MA
D000818:Animals; D001965:Bromides / Q000737:chemistry*; D002478:Cells, Cultured; D002499:Centrifugation, Density Gradient / Q000379:methods; D016054:DNA, Protozoan / Q000032:analysis; D033661:Oocysts / Q000166:cytology / Q000235:genetics / Q000302:isolation & purification / Q000648:ultrastructure; D009893:Opossums / Q000469:parasitology*; D017680:Potassium Compounds / Q000737:chemistry*; D011419:Propidium / Q000737:chemistry; D019105:Random Amplified Polymorphic DNA Technique; D012522:Sarcocystis / Q000254:growth & development / Q000302:isolation & purification*
D001965:Bromides; D016054:DNA, Protozoan; D017680:Potassium Compounds; D011419:Propidium; C039004:potassium bromide