Last month, the Parenteral Drug Association's 2015 Annual Meeting was held in Las Vegas with the central theme of “Manufacturing Innovation and Efficiency: Achieving Quality Performance in Sterile and Biopharmaceutical Operations.” The meeting featured three main tracks to address industry challenges:
- Improving Manufacturing Performance
- Changing Regulatory Landscape
- Technology, Advances and Expectations
Andrew Sage, PhD and Manager, Microbiology Applications at Rapid Micro Biosystems, presented his poster, “Optimizing bioburden testing of mammalian cell culture samples through the use of differential lysis, membrane filtration and analysis by a Rapid Micro Method (RMM),” which you can download here.
The one-step differential lysis method he presented is a superior way of preparing a mammalian cell culture for bioburden than traditional spread plating. The poster discusses the issues associated with the traditional method, the experiment’s results and the benefits of the new method.
Limitations of Current Methods
Sample preparation using traditional spread plating is both labor-intensive and time-consuming, as the 10 mL test volume suggested by regulatory guidance means microbiologists must analyze 20 plates (0.5 mL each). Standard bioburden and sterility devices are often limited to test volumes of 1 mL or less. Additionally, membrane filters can become clogged when cell culture concentration exceeds 5 x 106 cell/mL, and the traditional method requires incubation times ranging from 3 to 5 days.
With automated, growth-based rapid microbial methods that allow for a shorter time to result, a faster method of sample preparation for bioburden testing is needed to maximize the potential time saving.
Overview of the Optimized Method
With the one-step differential lysis method, the microbiologist samples up to 10 mL of fluid and adds the sample to the differential lysis reagent in a single-use septa vial. The cell culture sample is then filtered and tested using the Growth DirectTM System.
Figure 1. One-step differential lysis, membrane filtration and analysis by a Rapid Micro Method.
The poster outlines the details of the experiment, including the type of culture samples used, the comparison performed between the suggested method and the traditional testing.
While the poster outlines additional details, Figure 2 illustrates the recovery percentage of the one-step method in CHO and mouse murine cell lines, showing that the results obtained using the Growth DirectTM System were comparable to the controls and, in some cases, exhibited improved detection (>100% of the mean CFU of the control).
Figure 2. Recovery: % Detected in Growth DirectTM System vs. PBS titer and control *Acceptance criteria: 50-200% of control
Benefits of the Method
The method described in the study allows the suggested 10 mL sample volume to be tested without the need for labor intensive spread plating. The optimized method was more efficient, with a short filtration time of 3 minutes at a high cell concentration (1x108 cells in 10 mL), and comparable CFU detection results significantly faster than the 3 days required for the traditional method.
The results of the study show that the one-step lysis method used with the Growth DirectTM System for automated rapid detection can speed the time to result, so contaminants can be detected sooner.
Learn more about automated rapid detection and enumeration—get our free guide here.