Antisepsis, Disinfection, and Sterilization. Gerald E. McDonnell
Biological Indicator is used to monitor steam sterilization cycles. Reproduced with permission of 3M Health Care.
The carrier can consist of any material, with typical examples being paper, stainless steel, glass, and plastics. In its true definition, a biological indicator consists of the inoculated coupon placed into a “primary pack,” which can be a protective envelope or pouch, or within an assembled vial or ampoule. In their simplest form, biological indicators are present within a protective envelope, and following exposure to a given sterilization process, the inoculated coupon is aseptically removed from its pack and incubated in a specified growth medium to determine the presence or absence of spore viability. Due to the release of dipicolinic acid (see section 8.3.11) upon germination and outgrowth of the spores, pH indicator dyes can be used in the growth media to indicate the presence of viability before visible growth (turbidity) is observed. To minimize aseptic handling, self-contained biological indicators that include the inoculated carrier within a vial containing a sealed ampoule of growth medium have also been developed (Fig. 1.23). Following exposure, the medium ampoule is broken to allow incubation without handling of the coupon. Further “rapid-review” biological indicators are available that detect the presence of certain endospore enzymes (e.g., (α-D-glucosidase) whose destruction by heat correlates with the loss of viability of the spore; the presence of enzyme activity can be detected fluorimetrically and can give a rapid indication of spore viability (usually within 1 to 4 h). In addition, these indicators are further incubated to demonstrate the presence or absence of growth, as for traditional biological indicators. Various standards that define the requirements for and use of biological indicators are given in Table 1.22.
TABLE 1.21 Bacterial-endospore species used to monitor and validate sterilization processes
| Sterilization process | Biological indicator |
| Moist heat | Geobacillus stearothermophilus |
| Dry heat | Bacillus atrophaeus |
| Irradiation | Bacillus pumilus |
| Ethylene oxide | Bacillus atrophaeus |
| Low-temperature steam formaldehyde | G. stearothermophilus |
| Hydrogen peroxide vapor | G. stearothermophilus |
Chemical indicators change color (or provide another visible change) on exposure to a given disinfectant or sterilization process (Fig. 1.24). They can range from simple process indicators that indicate exposure to a given process parameter (e.g., exposure to heat, but not necessarily at the right temperature and for the right amount of time) to more specific integrator indicators, which change color only on exposure to multiple variables (e.g., temperature and time for steam sterilization and concentration or time, temperature, and humidity for ethylene oxide sterilization). They can be classified in various ways, and an example is given in Table 1.23.
Chemical indicators are widely used, as they give an instant result and in some cases (as with some integrators) can be correlated to a biological indicator result. Applications include specific direct parameters that are required for disinfection or sterilization (e.g., verification of the presence of a minimal concentration of a biocidal formulation prior to use or that a range of conditions have been met in a sterilizer), but also other, indirect variables that are important to the efficacy of the process (e.g., Bowie-Dick tests are used to confirm the adequate removal of air in prevacuum-type steam sterilizers [see section 5.2]). Examples of various standards that define the requirements for and use of chemical indicators are given in Table 1.24.
TABLE 1.22 Examples of biological-indicator standards
| Referencea | Title | Summary |
| ISO 11138-1 | Sterilization of Health Care Products—Biological Indicators—Part 1: General Requirements | General requirements for production, labeling, test methods, and performance characteristics of biological indicator systems to be used in the validation and routine monitoring of sterilization processes |
| ISO 11138-2 | Sterilization of Health Care Products—Biological—Part 2: Biological Indicators for Ethylene Oxide Sterilization Processes | Specific requirements for biological indicators used for ethylene oxide sterilization, including test organism and performance criteria |
| ISO 11138-3 | Sterilization of Health Care Products—Biological Indicators—Part 3: Biological Indicators for Moist Heat Sterilization Processes | Specific requirements for biological indicators used for moist-heat (steam) sterilization, including test organisms and performance criteria |
| ISO 14161 | Sterilization of Health Care Products—Biological Indicators—Guidance for the Selection, Use and Interpretation of Results | Guidance for the selection, use, and interpretation of results of biological indicators used in the development, validation, and routine monitoring of sterilization processes |
| EN 866-1 | Biological Systems for Testing Sterilizers and Sterilization Processes. Part 1—General Requirements | General requirements for production, labeling, test methods, and performance characteristics of biological indicator systems to be used in the validation and routine monitoring of sterilization processes |
| USP XXIII | Biological Indicators—Resistance Performance Tests | Testing of the resistances and population of biological indicators |
| EP 5.1.2 | Biological Indicators of Sterilization | Requirements for biological indicators, including population and resistance |
aISO, International Standards Organization; EN, European Norm, from the CEN (European Committee for Standardization); USP, United States Pharmacopeia; EP, European Pharmacopoeia.
Other, miscellaneous indicators include mechanical indicators, such as gauges or sensors that measure temperature, concentration, pressure, time, etc., that are used to monitor various physical parameters during