Pharmaceutical manufacturers are constantly under pressure. Supply chain staff is needed to ensure prompt availability of raw materials; Production personnel are constantly looking for ways to improve efficiencies and output; and Management is busy making sure everything is functioning profitably. In the midst of all of these forces, the quality laboratories - including microbiology - face their own demands. In addition to ensuring that products meet all of the quality criteria, they are under significant pressure to provide these results as quickly as possible.
For most pharmaceutical manufacturers, a number of critical testing and control points occur during bioprocessing, including:
Microbiological-specific testing throughout these steps often includes bioburden, sterility, Mycoplasma, and purified water testing, among others. The advancement of Rapid Microbiological Methods (RMMs) and the application for pharmaceutical manufacturing is providing an opportunity for microbiology labs to move beyond the traditional media-based procedures, allowing for automation, faster turn-around times, operational cost savings, and increased accuracy, reproducibility and sensitivity.
Regulatory agencies continue to move forward with reviews and acceptance of RMMs. Recent updates to Validation Guidance include:
- PDA Technical Report #33, Evaluation, Validation and Implementation of New Microbiological Testing Methods (updated Oct. 2013)
- Ph. Eur. 5.1.6, Alternative Methods for Control of Microbiological Quality (in revision)
- USP <1223>, Validation of alternative microbiological methods (in revision)
With all of the focus on meeting regulatory, quality and operational demands, it can be difficult to determine which Rapid Microbiological Method would be the best fit for your laboratory.
There are several questions one should consider when evaluating implementation of a RMM:
- What standards or regulatory criteria does your testing need to meet?
- How many samples are you currently analyzing? How many samples would you like to analyze?
- What is your time-to-result with your current test method? How does this fit into your current laboratory workflow? How would improving the time-to-result effect your lab’s workflow?
- What training/support is needed for your current method? For your proposed method?
- What is your current cost/sample? What is the cost of labor associated with the analysis?
- What is the value of inventory awaiting the results (raw material or finished goods)? Are there inventory carrying costs associated with those products/materials?
The Bd FaCsMicroCountTM system
While flow cytometry has been used in research laboratories for over 50 years, the application of this technology for pharmaceutical manufacturing quality laboratories has only developed more recently. Along with the advancement of optical systems, reagents and software, primary drivers to pharmaceutical manufacturing application development have included throughput, automation and ease of use. For both qualitative and quantitative testing, users require multiple sample results per hour for different organism types (bacteria, yeast and mold), with continuous sample feeding and “rush” sample options. Furthermore, there is an expectation of an instrumented system to automatically add reagents, vortex/mix and analyze each sample. Additionally, systems should be relatively easy to use, with simplified routine maintenance requirements, so that scientists can focus on the resulting data and analysis.
Figure 1: Diff erentiation of Microorganisms Based on Intensity Plot Position- Raw Materials
- Purified Water
- In-Process Samples
- Stock Culture Maintenance
- Fermentation Enumeration
- Biological Indicator Testing
For the purposes of this article, we will focus on sharing portions of two successful applications in which the BD FACSMicroCount System has been proven to enhance microbiological testing in the pharmaceutical industry: Screening Purified/Process Water and Mycoplasma Fermentation.
Screening purified/process Water
Pharmaceutical manufacturers are well aware of the importance of water in their processes. The testing of pharmaceutical water is critical. Knowing the test results of purified or process waters in a fraction of the time can help provide insights into when problems are arising and reduce potential downtime or product concerns. A study was undertaken to show MicroCount performance in detecting and enumerating various levels of microbiological contamination in clean, pharmaceutical water and how it correlates to traditional media methods.
As the results (figure 2) show, the BD FACSMicroCount was able to accurately enumerate P. aeruginosa at various concentrations in filtered, deionized water with strong correlation to traditional methods. The total analysis time for the BD FACSMicroCount was less than 10 minutes compared to multiple days for the traditional method.
Figure 2: BD FACSMicroCount Analysis of Pseudomonas aeruginosa in Pharmaceutical WatersMycoplasma Fermentation
Mycoplasma batch loss is an important concern due to the unique issues that arise in Mycoplasma monitoring. Mycoplasma are notoriously slow growing, taking over twelve days for viable counts by traditional methods. There are multiple technologies that can be used to effectively measure Mycoplasma fermentation, each with their own features and benefits. While some rapid methods have proven to be faster than traditional methods, they are often prone to subjective results interpretation. Inaccurate counts at any stage can result in loss of time, production and costly media.
The BD FACSMicroCount System has been shown to provide an accurate result in a fraction of the time of traditional methods, while also demonstrating reproducibility, and good correlation with traditional methods.
As illustrated in Figure 3, the BD FACSMicroCount System was able to reproducibly enumerate Mycoplasma samples at different concentrations with an analysis time of 5-10 minutes versus the traditional 7-14 days.
Figure 3: Enumeration of Mycoplasma by the BD FACSMicroCount System
Conclusion
Rapid Microbiological Methods continue to gain acceptance because of the scientific and operational advantages that they can provide. While new methods are likely to arise, proven methods, such as the flow cytometry-based BD FACSMicroCount System are already being accepted - and used by pharmaceutical manufacturers - because of the benefits it can provide, including:
- Product Release Validation - Meets regulatory requirements; direct correlation with traditional methods
- In-Process Quality - Earlier bioburden results allow early intervention and reduce risk of compromising future product
- Convenience - Multiple sample types and test protocols run simultaneously
- Operational Cost Savings - Reduce inventory related costs
- Labor Cost Reduction - Easy to use workflow and walkaway automation enables flexibility of resources
For additional information, please visit: www.bd.com/microcount