The Ultimate Guide to Biological Indicators: What They Are, Why They're Crucial, and How to Choose the Right Ones

Ensuring sterility in pharmaceutical manufacturing and medical device production isn’t optional—it’s essential. At the core of any effective sterilization process are biological indicators (BIs), the gold standard for proving that sterilization methods work. But what exactly are they, why are they so critical, and how do you pick the right ones? This guide provides the answers you need. 

What Are Biological Indicators? 

Biological indicators are test systems containing highly resistant microorganisms, typically bacterial spores, designed to challenge sterilization processes. If these microorganisms are eliminated, your sterilization process is likely capable of inactivating less resistant bioburden organisms.  

• What’s Inside: BIs often include spores of Geobacillus stearothermophilus (used for steam and VH₂O₂ sterilization) or Bacillus atrophaeus (used for ethylene oxide and dry heat sterilization). 
• Why They Matter: Unlike chemical indicators and probes that measure physical parameters (temperature, pressure), BIs confirm the biological kill—proving that microorganisms can’t survive the sterilization process. 
• How They Work: BIs are exposed to the sterilization cycle, then incubated to check for growth. If no spores survive, the sterilization process is capable of inactivating highly resistant microorganisms.  

Why Are Biological Indicators Crucial? 

• BIs provide evidence that your sterilization process is capable of achieving the desired Sterility Assurance Level (SAL). 

• Regulatory bodies like the FDA and EMA require the use of BIs for sterilization validation. Standards such as ISO 11138 outline specific requirements for BIs.  

• Contaminated products can lead to infections, product recalls, and reputational damage. BIs help confirm sterility, reducing risks and ensuring safety. 

• Sterilization and biodecontamination processes require precise validation. BIs assist in effective cycle development and help ensure these methods consistently deliver reliable results. 

Types of Biological Indicators and When to Use Them 

Different sterilization methods call for specific types of BIs. Here’s a quick guide: 

• Microorganism: Geobacillus stearothermophilus 
• Use Case: Surgical instruments, pharmaceutical products, reusable medical devices, and pharmaceutical piping. 

• Microorganism: Bacillus atrophaeus 
• Use Case: Temperature-sensitive materials, plastics, and devices incompatible with high heat. 

• Microorganism: Bacillus atrophaeus 
• Use Case: Glassware, oils, powders, and other heat-resistant items. 

• Microorganism: Geobacillus stearothermophilus 
• Use Case: Isolators, cleanrooms, and bio-decontamination of surfaces. 

• Microorganism: Bacillus pumilus 
• Use Case: Single-use medical devices, disposable products, and pharmaceuticals. 

How to Choose the Right Biological Indicators 

Choosing the right BI ensures your process is effective and compliant. Here’s what to consider: 

• Use microorganisms designed to challenge your specific sterilization process. For example, steam sterilization requires Geobacillus stearothermophilus, while EO uses Bacillus atrophaeus. 

• Select BIs with microorganisms more resistant than the bioburden in the most difficult-to-sterilize location and resistant enough to validate the process.
  

• Spore Strips: Simple and effective for routine testing. 
• Self-Contained BIs: Convenient with built-in growth medium for simplified culturing and reduced risk of post-process contamination.
• Liquid Ampoules: Designed for use when sterilizing liquid products.
• Stainless Steel Discs: Ideal for surface testing in isolators or challenging locations. 

• Verify that your BIs meet standards like ISO 11138.  

• Look for BIs that integrate seamlessly into your workflow and provide reliable, easy-to-interpret results. 

Best Practices for Using Biological Indicators 

To get the most from your BIs, follow these best practices: 

• Place BIs in the most challenging locations (cold zones, dense loads, cannulas, tubing) to ensure a true test of the sterilization process. 

• Incorporate BIs into cycle development, validation, and routine monitoring to ensure consistent performance. 

• While BIs provide the biological kill proof, chemical indicators give quick visual feedback on physical conditions. 

• Record all results, investigate any failures, and keep detailed documentation for audits and ongoing compliance. 

• Regularly analyze BI data to refine your sterilization processes and address any issues proactively. 

 Conclusion 

Biological indicators are essential for validating sterilization processes, meeting regulatory requirements, and ensuring product safety. They provide proof that your methods work, protect patients, and support compliance. 

Partnering with Mesa Labs gives you access to trusted biological indicators, expert tools, and the support you need to simplify validation and maintain sterility assurance. With Mesa Labs, you can confidently optimize your sterilization processes and ensure consistent, reliable results.