title: Purification of turkey coronavirus by Sephacryl size-exclusion chromatography.
abstract: Sephacryl S-1000 size-exclusion chromatography was used to purify turkey coronavirus (TCoV) from infected turkey embryo. TCoV was propagated in the 22-day-old turkey embryos. Intestines and intestinal contents of infected embryos were harvested and homogenized. After low speed centrifugation, the supernatant was concentrated by ultracentrifugation through a cushion of 30 or 60% sucrose solution, or by ammonium sulfate precipitation. The purification methods included sucrose gradient and Sephacryl S-1000 size-exclusion chromatography. Ultracentrifugation through a cushion of 60% sucrose solution was better than the other two methods for concentration of TCoV from intestinal homogenate. The most effective method for purifying TCoV and removing extraneous materials was size-exclusion chromatography as analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. More spike-rich particles were observed in the sample purified by chromatography than those purified by sucrose gradient as examined by electron microscopy. Differentiation of turkey anti-TCoV antiserum from normal turkey serum was better achieved by ELISA plates coated with TCoV preparation purified by size-exclusion chromatography than that purified by sucrose density gradient. The results indicated that Sephacryl S-1000 chromatography was useful for purification of TCoV.
authors.affiliation: Department of Veterinary Pathobiology and Animal Disease Diagnostic Laboratory, Purdue University, West Lafayette, IN 47907-1175, USA.
authors.forename: C C
authors.initials: CC
authors.lastname: Loa
authors.forename: T L
authors.initials: TL
authors.lastname: Lin
authors.forename: C C
authors.initials: CC
authors.lastname: Wu
authors.forename: T A
authors.initials: TA
authors.lastname: Bryan
authors.forename: H L
authors.initials: HL
authors.lastname: Thacker
authors.forename: T
authors.initials: T
authors.lastname: Hooper
authors.forename: D
authors.initials: D
authors.lastname: Schrader
mesh_terms: D000818:Animals; D000956:Antigens, Viral / Q000032:analysis; D002499:Centrifugation, Density Gradient / Q000379:methods; D002850:Chromatography, Gel / Q000379:methods; D004752:Coronavirus, Turkey / Q000254:growth & development / Q000302:isolation & purification*; D004625:Embryo, Nonmammalian / Q000821:virology; D004753:Enteritis, Transmissible, of Turkeys / Q000821:virology; D006386:Hemagglutination Tests; D007422:Intestines / Q000821:virology; D014422:Turkeys; D014461:Ultracentrifugation / Q000379:methods
chemical_list: D000956:Antigens, Viral