Liquid Submersible Biological Indicators (LSBI) are designed to provide microbiological lethality measurements for the moist heat sterilization of containerized liquids. Containerized liquid products primarily include pharmaceuticals (contained in vials, ampoules or prefilled syringes), or other liquid components and lab materials. LSBIs can also be used for monitoring the sterilization of medical waste products which are often a mix of liquid and solid items.
The LSBIs design is quite similar to the product being sterilized in that they consist of containerized liquid. The liquid is a specially formulated culture medium containing surrogate test organism, hermetically sealed into glass ampoules. The BIs are produced according to relevant USP1 and ISO2,3 standards. Mesa offers a family of LSBIs including SterilAmp®, MagnaAmp®, ProSpore®, and SterilAmp® 5230 (Figure 1).
Figure 1. Liquid Submersible Biological Indicators formats and size comparisons
These BIs configurations are all quite similar in their design and function but are varied enough to meet each manufacturer’s specific requirements. A basic description of each BI is presented in Table 1. Additional technical information (e.g.; Product Specification, Technical Report, Population Assay Instructions) are provided at mesalabs.com.
LSBIs are used throughout the sterilization life cycle including during process development, validation, and routine monitoring. The appropriate LSBI for the sterilization cycle is determined in the process development stage and will depend on the validation approach (e.g. Overkill, Bioburden/Biological Indicator).
It is important that the LSBIs are only used to monitor the moist heat sterilization of containerized liquid loads and not dry or porous items. LSBI are not sensitive to air pockets that may exist in these loads and give a false indication of lethality.
When sterilizing liquids in containers, the steam does not come into direct contact with the product within. Rather, steam delivers its energy to the container, which then indirectly delivers it to the liquid by conduction. The bioburden organisms present in the liquid are then inactivated as lethality increases in the product. Contrast this to moist heat sterilization of hard or porous items where thermal energy is directly delivered to the surface of the product and the organisms thereon.
LSBIs are designed to be suspended in the liquid product inside the container. The slowest to heat area in containerized liquids is typically in the geometric center slightly above the bottom of the container. In larger containers, this will be the area of lowest lethality and is the recommended location to suspend the Bl.
The volume of liquid inside the containers being sterilized will determine which LSBI is most appropriate. For volumes of liquid ≥100 ml, MagnaAmp and ProSpore are the recommended BIs. These BIs have a "neck" around which a fine gauge wire can be easily attached to suspend the ampoule in the liquid where it can monitor this most difficult to sterilize location.
For volumes of liquid less than 100 ml, SterilAmp is the recommended BI. Smaller volumes of containerized product will heat more consistently throughout with minimal lag time in the slower to heat areas. SterilAmp is small in size (26 x 6.5 mm or 18 x 6.5 mm) and will easily fit inside most smaller containers.
In some instances, the liquid Bl may not need to be placed into the liquid but rather can be placed at various locations in the autoclave next to the containers of liquid. Generally, MagnaAmp or ProSpore are used in this manner when sterilizing vials of product that are approximately the same volume and with a known similar resistance as the MagnaAmp or ProSpore BI units. Used in this manner, the Bl will function as a surrogate vial and will experience similar conditions as the product vials.
The SterilAmp 5230 LSBI is used for heat sensitive products that are processed using the Bioburden/Biological Indicator validation approach. These processes deliver less thermal insult to the product (as compared to the Overkill method) while still demonstrating an acceptable Sterility Assurance Level (SAL). This is accomplished by basing the SAL on the bioburden while validating the process using the surrogate test organism Bacillus subtilis. B. subtilis is significantly more resistant than the typical bioburden, but not as resistant as the more commonly used organism, Geobacillus stearothermophilus.
The positions of the LSBI in a loaded sterilizer should be based on thermocouple profiling of the loaded chamber to assure that the "most difficult to sterilize" locations are being monitored. Additionally, BIs are also typically located from top to bottom, front to back, and in the geometric center of the load.
LSBIs are recommended when sterilizing biohazardous waste. This waste is often a combination of liquids and solids and the use of spore strips or self-contained BIs is not appropriate as these can lead to false positive results. A common practice is to embed the LSBIs in the load in the slow to heat areas. For easy retrieval of the BI unit post sterilization, it is recommended that a wire be placed around the neck of the LSBI (leaving the other end of the wire outside of the bag).
Mesa Labs has a complete LSBI portfolio with units ranging in size, and resistance to the moist heat sterilization process. This allows the users to select the appropriate BI for their product and selected validation approach. LSBI are easy to use with no post processing manipulations (only an incubator is needed) and most have results within 48 hours.
Accessory Devices
Small table top incubators are available for incubation of the MagnaAmp (Model 1510) and ProSpore (1480) LSBIs (see image below). These units are preprogrammed to the optimal incubation temperature and will hold 15 and 11 BI units respectively.
References
1. USP 1229.5 Biological Indicators for Sterilization
2. ISO 11138–1: 2017 Sterilization of health care products—Biological indicators—Part 1: General requirements
3. ISO 11138–3: 2017 Sterilization of health care products—Biological indicators—Part 3: Biological indicators for moist
heat sterilization processes
SN092-V1