title: Systematic evaluation of in vitro and in vivo adventitious virus assays for the detection of viral contamination of cell banks and biological products.
abstract: Viral vaccines and the cell substrates used to manufacture them are subjected to tests for adventitious agents, including viruses, contaminate. Some of the compendial methods (in vivo and in vitro in cell culture) were established in the mid-20th century. These methods have not been subjected to current assay validation, as new methods would need to be. This study was undertaken to provide insight into the breadth (selectivity) and sensitivity (limit of detection) of the routine methods, two such validation parameters. Sixteen viral stocks were prepared and characterized. These stocks were tested in serial dilutions by the routine methods to establish which viruses were detected by which methods and above what limit of detection. Sixteen out of sixteen viruses were detected in vitro, though one (bovine viral diarrhea virus) required special conditions to detect and another (rubella virus) was detected with low sensitivity. Many were detected at levels below 1 TCID50 or PFU (titers were established on the production cell line in most cases). In contrast, in vivo, only 6/11 viruses were detected, and 4 of these were detected only at amounts one or more logs above 1 TCID50 or PFU. Only influenza virus and vesicular stomatitis virus were detected at lower amounts in vivo than in vitro. Given the call to reduce, refine, or replace (3Rs) the use of animals in product safety testing and the emergence of new technologies for the detection of viruses, a re-examination of the current adventitious virus testing strategies seems warranted. Suggested pathways forward are offered.
authors.affiliation: Charles River Laboratories, 358 Technology Drive, Malvern, PA 19355, United States.
authors.forename: James
authors.initials: J
authors.lastname: Gombold
authors.affiliation: Charles River Laboratories, 358 Technology Drive, Malvern, PA 19355, United States.
authors.forename: Stephen
authors.initials: S
authors.lastname: Karakasidis
authors.affiliation: Charles River Laboratories, 251 Ballardvale St. Wilmington, MA 01887, United States.
authors.forename: Paula
authors.initials: P
authors.lastname: Niksa
authors.affiliation: Charles River Laboratories, 358 Technology Drive, Malvern, PA 19355, United States.
authors.forename: John
authors.initials: J
authors.lastname: Podczasy
authors.affiliation: Charles River Laboratories, 358 Technology Drive, Malvern, PA 19355, United States.
authors.forename: Kitti
authors.initials: K
authors.lastname: Neumann
authors.affiliation: Advanced BioScience Laboratories, 9800 Medical Center Dr. Bldg. D, Rockville, MD 20850, United States.
authors.forename: James
authors.initials: J
authors.lastname: Richardson
authors.affiliation: Advanced BioScience Laboratories, 9800 Medical Center Dr. Bldg. D, Rockville, MD 20850, United States.
authors.forename: Nandini
authors.initials: N
authors.lastname: Sane
authors.affiliation: Advanced BioScience Laboratories, 9800 Medical Center Dr. Bldg. D, Rockville, MD 20850, United States.
authors.forename: Renita
authors.initials: R
authors.lastname: Johnson-Leva
authors.affiliation: Wyeth, 401N Middletown Rd., Pearl River, NY 10965, United States.
authors.forename: Valerie
authors.initials: V
authors.lastname: Randolph
authors.affiliation: Crucell, Newtonweg 1, 2333 CP Leiden, PO Box 2048, 2301 CA Leiden, The Netherlands.
authors.forename: Jerald
authors.initials: J
authors.lastname: Sadoff
authors.affiliation: National Institute for Biologics Standards and Control, Blanche Lane, South Mimms, Potters Bar, Hertfordshire EN6 3QG, United Kingdom.
authors.forename: Phillip
authors.initials: P
authors.lastname: Minor
authors.affiliation: GSK Vaccines, Rue de l'Insitut 89, 1330 Rixensart, Belgium (formerly NIH/NIAID).
authors.forename: Alexander
authors.initials: A
authors.lastname: Schmidt
authors.affiliation: Merck and Co., Inc., WP17-101, 770 Sumneytown Pike, P.O. Box 4, West Point, PA 19486, United States.
authors.forename: Paul
authors.initials: P
authors.lastname: Duncan
authors.affiliation: NIH/NIAID Division of AIDS, 6700B Rockledge Dr., Rm. 5145, Bethesda, MD 20892, United States. Electronic address: rsheets@grimalkinpartners.com.
authors.forename: Rebecca L
authors.initials: RL
authors.lastname: Sheets
mesh_terms: D000818:Animals; D001688:Biological Products / Q000032:analysis* / Q000592:standards; D002460:Cell Line; D002645:Chickens; D004340:Drug Contamination / Q000517:prevention & control*; D006801:Humans; D057230:Limit of Detection; D051379:Mice; D010063:Ovum; D011786:Quality Control; D014765:Viral Vaccines / Q000592:standards; D014780:Viruses / Q000302:isolation & purification*
chemical_list: D001688:Biological Products; D014765:Viral Vaccines
keywords: 3Rs; Adventitious agents; In vitro; In vivo; Vaccines; Viral safety