Key Benefits and Challenges of In-House Propagation of Reference Cultures
Reference Cultures related Best practices
BEST PRACTICES FOR REFERENCE CULTURES
Key Benefits and Challenges of In-House Propagation of Reference Cultures
In-House Culture Propagation (The "Seed Lot" System)
Many laboratories across the globe are still following an in-house culture propagation method. As a standard practice, they acquire/purchase an original reference strain (s) from a culture collection and then prepare their own long-term frozen stocks and working cultures. Here in this post, we will evaluate both benefits and challenges related to this practice.
Benefits
Low Cost (per test): Once the initial reference strain is purchased, a lab can maintain long term frozen stock or mother culture to produce a large number of working cultures, significantly lowering the cost per test over time.
Flexibility and Availability: An in-house collection provides immediate access to QC strains, which can be useful for labs that frequently perform tests or have a large volume of QC.
Complete Control: The lab has full control over the preparation, storage, and passage of its QC strains, which can be advantageous for research or specialized applications where specific conditions are required.
Challenges
Specialized Expertise and Labour: Preparing and maintaining a seed lot system is labour-intensive and requires specialized expertise techniques, strict aseptic techniques, adequate cryo-preservation methodology, and meticulous record-keeping (for internal and external audits).
Risk of Genetic Drift (Over-Passaging): The greatest risk of in-house propagation is exceeding the passage-limit. Subculturing the reference strain beyond certain passage (such as beyond 05th Passage as per USP) increases the chance of genetic drift/mutations, which can lead to changes in the organism's phenotypic characteristics (e.g., an S. aureus losing its methicillin resistance). Therefore, stick with passage capping limit to mitigate this risk.
Risk of Contamination: In-house propagation required repeated manual handling and subculturing, which throw the other challenge that is a higher risk of introducing a contaminating organism due to manual error.
Equipment and Maintenance Costs: Maintaining a long-term frozen stock requires a -70°C or -80°C deep-freezer, which requires purchase cost of the deep freezer, cost for maintenance, alarm system for doors, UPS back-up etc. and time to time disinfection of it to stay compliant.
Complex Documentation and Auditing: Meticulous records must be kept for each passage, including date of purchase of reference strains, media, and technician, to demonstrate compliance. This can be complex and time-consuming during audits.
Variability: Despite best efforts, there can be subtle variations in culture preparation and handling that lead to more variability in test results compared to standardized commercial products.
Best Practices
Regardless of the method chosen, key best practices include:
Documentation: Maintain meticulous records of all QC activities, including the source, lot number, expiration date, and performance of each QC organism.
Traceability: Ensure all QC strains are traceable to a reputable national culture collection.
Adherence to Guidelines: Strictly follow CLSI and EUCAST guidelines for QC frequency and performance limits.
Proper Storage: Store QC materials according to the manufacturer's or in-house protocol to maintain viability and prevent degradation. For in-house stocks, use appropriate cryoprotectants and a reliable ultra-low temperature freezer.
Regular Verification: Periodically, verify the identity and expected characteristics of in-house cultures to ensure they have not mutated.