O-033. Application for a Rapid Flow Cytometry Method for Process Control Bioburden Testing in Live Attenuated Influenza Vaccine Intermediates

P. K. Bhusari1, M. S. Galinski1, A. M. Steger2;
1MedImmune Vaccines, Inc., Mountain View, CA, 2Advanced Analytical Technologies, Inc., Ames, IA.

Embryonated chicken eggs used in influenza vaccine manufacturing, although are specific pathogen free (SPF), often contain microorganisms. Microorganisms, when present at unacceptable levels of bioburden, can interfere with vaccine production. To minimize the level of bioburden in raw vaccine bulk, allantoic fluids from the eggs are harvested into multiple sub-lots which are then screened for the presence of bioburden. Highly contaminated sub-lots are discarded prior to downstream processing, which subsequently removes all remaining bioburden. This process warrants the use of a rapid bioburden detection method that provides a quick assessment of bioburden levels to facilitate straight-through processing, without having to quarantine the in-process bulk. The ability of the Micro PRO™ flow cytometer (Advanced Analytical, Ames, IA) to address this issue was demonstrated. Sub-lots of vaccine material from a mock production run were analyzed on the Micro PRO™ system using Advanced Analytical’s FASTEST Total Viable Organisms detection kit. Prior to testing samples on the Micro PRO™ detection system, samples were pre-treated using a novel method comprising of filter clarification using 0.45μm cellulose acetate membrane filters and a detergent wash to reduce the background noise contributed by cellular debris and other components of allantoic fluid. All samples were plated in parallel on Tryptic Soy Agar plates for count comparisons. Eleven percent of the sub-lots were positive for bioburden with counts ranging from 104 to 105 cfu/ml. The Micro PRO™ system provided accurate detection of relative contamination levels in each of the contaminated sub-lots of vaccine material. This rapid bioburden technique provides a faster time to result than the conventional bioburden method and increases the reliability of vaccine production.

245/O. Antibiotics, Antimicrobials and Mycotoxins

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