Downstream Processing: A Revalidation Study of Viral Clearance in the Purification of Monoclonal Antibody CB.Hep-1 - - BioPharm International

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Downstream Processing: A Revalidation Study of Viral Clearance in the Purification of Monoclonal Antibody CB.Hep-1

To increase the inactivation factor of the whole sanitization procedure, the resistance of nonenveloped viruses HPV-2 and CPV was also studied in 0.1 N HCl (pH 1.0). The isoelectric pH of the purified poliovirus and CPV ranges from 5.5 to 7.0.24 This attribute will be drastically changed under the presence of the 0.1 N HCl allowing, in theory, a high virus inactivation factor. Results demonstrated this. HPV-2 was inactivated 6.13 Logs in 7 h and CPV was inactivated 6.60 Logs in just 1 h. Therefore, a robust sanitization protocol for the affinity chromatography step of a MAb CB.Hep-1 purification process should involve the combination of 70% ethanol and 0.1 N HCl in the studied exposure time.

CONCLUSIONS

A scale down of the MAb CB.Hep-1 purification process demonstrated removal factors of enveloped and nonenveloped model viruses similar to the first validation study. HSV-1, HIV-1, and CPV are sensitive to incubation with 70% ethanol (from 3.0 to 4.6 Logs). We find that 0.1 N HCl provides robust inactivation of more than 6.13 Logs of nonenveloped high resistance viruses. Ethanol at 20% concentration inactivated 3.7 Logs of enveloped viruses HSV-1 and HIV-1 but was totally ineffective in inactivating the nonenveloped viruses HPV-1 and CPV.

ACKNOWLEDGMENTS

The authors kindly thank the Cuban National Center for Animal Breeding for producing and supplying the specific pathogen-free ascitic fluid used in this work.

Rodolfo Valdés is head of the monoclonal antibody production department at the Center for Genetic Engineering and Biotechnology (CGEB), Havana, Cuba, 537.2716022, ext. 7101,

María del Rosario Alemán is the head of the cell culture laboratory of the monoclonal antibody production department at CGEB.

Andrés Tamayo, Sigifredo Padilla, Lamay Dorta, and Biunayki Reyes are researchers at the monoclonal antibody production department at CGEB.

Enrique Noa, Marta Dubed, and Giselle Alvarez are researchers at the National Reference Center for AIDS Research, Havana, Cuba.

Déborah Geada and Leonor Navea is a researcher at the Research Institute of Tobacco, Department of Industrial Development, Havana, Cuba.

Tatiana González is an engineer at the Process Control Department of CGEB. Eutimio Gustavo Fernández is a researcher at Inorganic Chemistry Department, Center for Engineering and Chemical Researches, Havana, Cuba.

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