1025-028X 1025-028X S1025-028X2000000200005 Cuba 00 06 2000 00 06 2000 9 2 23 28

ARTICULOS ORIGINALES

 

 

Ensayo in vivo para determinar agentes extraños en células BHK-21 empleadas en la obtención de biológicos.

 

In vivo determination of adventitious agents in BHK-21 cells used in the production of
biologicals.

 

]]> Georgina Pardo, Ernesto Almora, Odalys Fidalgo, Arelys Zamora, Niurka Rodríguez y Ela María Pérez.

Instituto Finlay, Centro de Investigación-Producción de Vacunas y Sueros. Ciudad de La Habana, Cuba. E-mail:
gpardo@finlay.edu.cu

RESUMEN

La controversia sobre los parámetros de calidad de las células de línea como substrato para la obtención de
productos farmacéuticos ha sido un tema de interés desde 1987. Las pruebas realizadas a las células, generalmente siguen las recomendaciones de los “Puntos a considerar en la Caracterización de Líneas Celulares empleadas para la Producción de Biológicos” entre las que se encuentra que estén libres de agentes adventicios. En este estudio al clon 13 de células BHK-21 procedente de riñón de hámster sirio recién nacido, se le determina la presencia de agentes extraños in vivo; así como su inocuidad en animales de experimentación. La prueba in vivo incluyó la inoculación de embriones de pollo por cavidad alantoidea y saco vitelino, ratones lactantes y adultos, curieles y conejos. Todos los animales fueron observados para determinar signos clínicos de infección viral. En los embriones se observó la viabilidad. En todos los casos se mantuvo el 100% de los embriones y animales vivos y saludables durante el período de observación. Las células del banco estaban libres de virus hemaglutinantes y hemadsorbentes, virus de la coriomeningitis linfocítica y coxsackie. Los pesos de los animales inoculados fueron mayores que los de los animales controles. Las células no presentaron evidencia de agentes extraños en los ensayos in vivo y fueron bien toleradas e inocuas en los animales de experimentación.

Palabras claves: Caracterización, células BHK-21, agentes extraños, animales de experimentación

ABSTRACT

]]> Acceptance of cell lines as a substrate for production of pharmaceuticals has been a controversial topic since 1987. The cells are generally tested according to the recommendations that appear in “Points to Consider in the
Characterization of Cell Lines Used to Produce Biologicals”. They should be free from adventitious agents; among others in vitro and in vivo tests are recommended for this purpose. In this report BHK-21-cell clone 13 from new born Syrian hamster kidney cells is assayed in vivo to determine adventitious agents; it is also tested for nonspecific innocuousness in experimental animals. In vivo tests included the inoculation of adult mice,suckling mice, guinea pigs, rabbits, and embryonated chicken eggs by the yolk sac and allantoic routes of inoculation. All animals were observed for clinical signs of viral infection. Egg viability was determined. The results showed that 100% of the animals and eggs remained healthy and survived the observation period. The cell bank was free of haemadsorbent, haemagglutinating, lymphocytic chroriomeningitis and Coxsakie viruses. The weights of the tested animals were higher than the weights of the control animals. The cells did not present evidences of adventitious agents in the in vivo assays and they were well accepted by the experimental animals.

Key words: Characterization, BHK-21 cells, adventitious agents, experimental animals.

texto completo en pdf

REFERENCIAS

1. WHO. Technical Report Series (TRS), Annex 1. Requirements for Biological Substances No. 50. Final

2. Draft. Requirements for use of animal cells as in vitro substrates for the production of biological. Geneva: WHO;1997.

3. FDA. Points to consider in the characterization of cell lines used to produce biologicals. USA:FDA; 1993.

4. European Pharmacopoeia. Cell Culture for the production of Veterinary Vaccines; 1998.

5. Macpherson I. Stocker,M. Polyoma transformation of hámster cell clones- an investiagation of genetic factor affecting cell competence. Virology. 1962; 16:147-151

6. Wiktor TJ, Sokol F, Kuwert E y Koprowski H. Proc Soc Exp Biol (NY) 1969; 131:799.

7. Kouri J, Kouri G, Bravo E, Rodríguez P and Aguilera A. Ultrastructure of BHK-21 cells as revealed by freeze-etching and fixation methods. J Microscopie 1971; 11:331-338.

8. Sureau P. Rabies vaccine production in animal cell culture. Advances in biochemical engineering and biotechnology 1987; 34:11-128.

9. Hay RJ, Caputo J y Macy ML. ATCC Quality Control Methods for Cell Lines 1992; 132 pp.

10. Kamrud KI, Powers AM, Higgs S., et al. The expression of chloramphenicol acetyltransferase in mosquitoes and mosquito cells using a packaged Sindbis replicon system. Exp-Parasitol 1995; 81(3):394-403.

11. Nicholson KG. Cell culture vaccines for human use: General considerations. En: Meslin FX., Kaplan MM. and Koprowski H. Laboratory techniques in rabies. Fourth Edition. Geneva:WHO. 1996:271-279.

12. Bieniasz PD, Erlwein O, Aguzzi A, Rethwilm A, McClure MO. Gene transfer using replicationdefective human foamy virus vectors. Virology. 1997; 235(1):65-72.

13. Capstick PB, et al. Growth of a clone strain of hamster kidney cells in suspended cultures and their susceptibility to the virus foot-and-mouth disease. Nature 1962; 195 (4847):1163-1164.

14. Reculard P. Cell culture vaccines for veterinary use. En Meslin FX, Kaplan MM and Koprowski H. Laboratory techniques in rabies: Fourth Edition. Geneva:WHO; 1996; 314-322.

15. Wilde H. Rabies 1996. Int J Infect Dis. 1997; 1:135-1429.

16. Montes de Oca H. Higk Yield Method for Kidney Tissue. En: Kruse PF, JR and MK Patterson, J R.eds. Tissue Culture Methods and Aplications.1973; 8-12.

17. Guías para el cuidado y uso de animales de laboratorio; 1996.

18. Jawetz E, Melnick J L, Adelberg E A. Manual de Microbiología Médica. 9ª Edición 1985; 595 pp.NC-26-67-83. Medicamentos. Prueba de inocuidad. Método de Control.

19. Lee CK. Issues of Biological Assays for Viral Vaccines. Dev Biol Stand 1996; 88: 41-47.

20. IFFA CREDO: Animaux de Laboratoire. 1986; 72-73.

 

]]> WHO 1997 FDA 1993 European Pharmacopoeia 1998 1962 16 147-151 1969 131 799 1971 11 331-338 1987 34 11-128 1992 132 1995 81 3 3 394-403 1996 Fourth 271-279 1997 235 1 1 65-72 1962 195 4847 4847 1163-1164 1996 Fourth 314-322 1997 1 135-1429 1973 8-12 1996 1996 88 41-47 IFFA CREDO 1986 72-73