Thermal Methods
Thermal sterilization is the most common approach. Its efficacy depends on temperature: while vegetative bacterial forms die at 60 °C, spores, owing to their dense coats and bound water, are inactivated only at 160–170 °C.
- Dry heat. Performed in dry-heat sterilizers (hot-air ovens, Pasteur ovens). Standard regimen: 180 °C for 60 minutes (alternatively, 160 °C for 150 minutes). Only thermostable objects are processed: laboratory glassware, metal instruments, and silicone rubber items.
- Steam under pressure (autoclaving). The physical principle is that increasing pressure inside the autoclave shifts the boiling point of water upward. At a pressure of 2 atmospheres, steam temperature reaches 121 °C. The standard regimen (121 °C for 15–20 minutes) reliably kills spores. The method is used for dressings, linens, culture media, solutions, and the disposal of infectious material. Interestingly, hyperthermophilic archaebacteria can reproduce at 100 °C and even survive an hour of standard autoclaving.
Fractional Sterilization (Tyndallization)
This method is a lifesaver for thermolabile materials (e.g., culture media containing carbohydrates) that are irreversibly destroyed at temperatures above 100 °C.
The process spans three days. The object is heated daily to 70–80 °C for 30–60 minutes. The first heating destroys vegetative microbial cells. During the intervals between heating sessions, the object is placed in an incubator: this prompts surviving spores to germinate and transform into vulnerable vegetative forms. Repeated heating on subsequent days completely destroys this "new wave" of microorganisms.
Chemical and Gas Processing
- Gas sterilization. Utilizes toxic gases such as ethylene oxide, an ethylene oxide-methyl bromide mixture, or formaldehyde. Gases act as alkylating agents, inactivating proteins and enzymes and damaging nucleic acids. Processing takes place in hermetic chambers at 20–60 °C in the presence of moisture. Due to the high penetrating power of gases, the method is ideal for optics, polymers, electronics, and protein media.
- Endoscope processing. Complex equipment made of thermolabile materials is subjected to immersion. They are submerged in solutions of chemical sterilants, most commonly formaldehyde or glutaraldehyde. After sterilization, the device must be rinsed of the solution, dried, and stored under strict aseptic conditions for no more than 3 days.
Physical Methods
- Radiation sterilization. Widely used in industry for processing massive batches of single-use medical devices (syringes, blood transfusion sets). The active agent is a beam of accelerated electrons or $\gamma$-radiation from radioactive isotopes. Radiation fatally damages microbial nucleic acids.
- Filtration sterilization (mechanical method). Used to free thermolabile liquids from microorganisms: blood serum, drug solutions, and culture media. The liquid is passed through ceramic, asbestos, or glass filters. Membrane nitrocellulose filters are considered the "gold standard" in this group.
Promising technologies also include sterilization using ozone and ionized plasma.
Efficacy Monitoring
Four groups of methods are used to verify equipment performance:
- Microbiological. Inoculation of a portion of the sterilized object onto nutrient media (aerobes, anaerobes, fungi). The method is highly precise, but due to its duration, it is not used in routine practice.
- Chemical. Use of indicators—substances that change color or physical state at the required temperature (ampoules with benzoic acid or urea, indicator strips). They are placed both on the surface and deep inside objects.
- Biological (biotests). Use of spores from highly resistant bacteria (Bacillus stearothermophilus, Bacillus licheniformis). Spores are hidden inside items; if they perish, the cycle was effective.
- Technical. Regular inspection of equipment functionality by specialized engineering services.