1. What is an autoclave and what is its principle?
An autoclave is a steam sterilization device used to destroy microorganisms using high-pressure saturated steam.
The principle is moist heat sterilization, where saturated steam condenses on the surface, releasing latent heat and killing microorganisms rapidly.
Operates on steam penetration, heat transfer, and pressure-temperature relationship.
2. What is the difference between dry heat sterilization and autoclave sterilization?
Dry heat uses hot air with no moisture and requires higher temperatures and longer exposure.
Autoclave sterilization uses saturated steam, which transfers heat more efficiently.
Autoclave is faster, more effective, and suitable for most pharma and chemical equipment.
3. What is the working principle of steam sterilization?
Steam condenses on cooler surfaces and releases latent heat.
Moist heat denatures proteins and destroys microbes including spores.
Sterilization effectiveness depends on temperature, pressure, time, and steam quality.
4. What are the standard autoclave operating parameters (temperature, pressure, time)?
Common sterilization conditions are 121°C at 15 psi (1.1 bar) for 15–30 minutes.
For rapid cycles, 134°C at 2.1 bar is used for short exposure.
Parameters depend on load type, volume, and bioburden.
5. What is the importance of saturated steam in autoclaving?
Saturated steam carries maximum latent heat for effective sterilization.
Ensures rapid heat transfer and uniform temperature throughout the load.
Provides predictable and reliable microbial kill.
6. What is the difference between saturated steam and superheated steam?
Saturated steam contains moisture and condenses on surfaces, releasing latent heat.
Superheated steam behaves like dry gas and does not condense easily.
Superheated steam is ineffective for sterilization.
7. Why is superheated steam not suitable for sterilization?
It lacks moisture required to transfer latent heat.
Does not condense on the load surface, resulting in poor microbial kill.
Can cause overheating without effective sterilization.
8. What is F₀ value?
F₀ is the equivalent sterilization time at 121.1°C that provides a specific microbial lethality.
Used to quantify sterilization effectiveness.
Higher F₀ indicates greater microbial reduction.
9. How do you calculate F₀ value?
F₀ is calculated using the time–temperature integration method based on Z-value.
Formula: F₀ = ∫ 10^((T – 121.1)/Z) dt
Where T is temperature in °C and Z is the temperature change required for tenfold reduction in D-value.
10. What is D-value and Z-value in sterilization?
D-value is the time required to achieve 90% reduction (1 log) of a specific microorganism at a specific temperature.
Z-value is the temperature change needed to change the D-value by one log.
These values are used to design sterilization cycles and lethality calculations.
11. What is Bowie-Dick test and why is it performed?
The Bowie-Dick test checks the efficiency of air removal and steam penetration in pre-vacuum autoclaves.
Ensures that no air pockets remain, which would prevent proper sterilization.
Performed daily before sterilization cycles as a safety and validation requirement.
12. What is the purpose of a vacuum leak test in autoclaves?
It verifies the integrity of the chamber and piping by checking for unwanted air ingress.
Ensures that the vacuum system can hold the required vacuum level for a set duration.
Detects gasket leaks, valve leaks, or mechanical faults.
13. What is pre-vacuum and post-vacuum in autoclave cycle?
Pre-vacuum removes trapped air from the load and chamber to ensure full steam penetration.
Post-vacuum removes residual steam at the end of the cycle for effective drying.
These steps improve sterilization reliability and load dryness.
14. What is air removal efficiency and why is it important?
Air removal efficiency indicates how effectively the autoclave eliminates air from the chamber.
Air inhibits steam contact and prevents proper condensation and heat transfer.
High air removal efficiency ensures uniform sterilization across the load.
15. Why is air considered the enemy of steam sterilization?
Air creates cold spots by reducing steam temperature.
Prevents steam from contacting surfaces and transferring latent heat.
Leads to incomplete sterilization and failed cycles.
16. What are the main components of an autoclave?
Chamber, jacket, steam generator or steam supply, vacuum pump, steam trap, sensors, control system, safety valves, and door locking mechanism.
Each component supports steam generation, distribution, monitoring, and safety.
17. What is the role of a steam trap in autoclave?
A steam trap removes condensate while retaining steam.
Prevents waterlogging in steam lines, ensuring only dry saturated steam enters the chamber.
Improves steam quality and sterilization efficiency.
18. What is the purpose of a drain strainer in autoclaves?
Prevents debris or particles from clogging the drain line.
Ensures proper condensate removal and avoids chamber flooding.
Supports uniform temperature distribution at the chamber base.
19. What is the condensate drain function?
Removes condensed steam continuously during the cycle.
Helps maintain saturated steam conditions inside the chamber.
Ensures proper heating and eliminates cold spots.
20. What are the different types of autoclaves?
Gravity displacement autoclaves for basic sterilization of non-porous loads.
Pre-vacuum (vacuum-assisted) autoclaves for porous or complex loads.
Horizontal and vertical autoclaves based on chamber orientation.
Industrial, GMP-grade, and laboratory autoclaves depending on application.
21. What is a horizontal vs vertical autoclave?
A horizontal autoclave has a front-loading door and larger chamber suitable for industrial and pharma loads.
A vertical autoclave is top-loading, compact, and used for small laboratory loads.
Horizontal units offer better ergonomics and higher capacity.
22. What is a vacuum-assisted autoclave?
A vacuum-assisted autoclave uses a vacuum pump to remove air before steam entry.
Ensures steam penetrates porous loads, garments, and wrapped items.
Provides faster come-up time and more reliable sterilization.
23. What is a gravity displacement autoclave?
A gravity autoclave relies on steam entering from the top and pushing air downward through the drain.
Suitable for non-porous loads like glassware and metals.
Less effective for porous or packaged loads due to limited air removal.
24. What is the purpose of load temperature probes?
Load probes measure the actual product or load temperature during sterilization.
Ensure that the coldest spot reaches sterilization temperature.
Used for validation and routine cycle monitoring.
25. What is chamber temperature vs load temperature?
Chamber temperature is the air–steam mixture temperature inside the autoclave.
Load temperature is the actual temperature inside the product or items being sterilized.
Load temperature is more critical for confirming sterilization.
26. What is equilibrium time in autoclave?
Equilibrium time is the duration required for the load and chamber temperatures to match within defined limits.
Indicates that steady and uniform heating has been achieved.
Starts when the slowest heating point reaches target temperature.
27. Why is hold time required?
Hold time ensures the load remains at sterilization temperature long enough to achieve lethality.
Compensates for thermal lag in dense or porous loads.
Forms the core sterilization phase based on F₀ requirements.
28. What is the sterilization cycle sequence?
Typical sequence includes pre-vacuum, steam admission, come-up time, hold time, post-vacuum, and drying.
Each step ensures proper air removal, steam penetration, and load drying.
Sequence may vary depending on load type and cycle design.
29. What is cycle validation of autoclave?
Validation confirms the autoclave consistently achieves sterilization under defined conditions.
Includes heat distribution, heat penetration, and biological indicator tests.
Required to meet GMP, FDA, and quality standards.
30. What IQ, OQ, PQ tests are done for autoclave?
IQ verifies installation, utilities, materials, and documentation.
OQ verifies functional performance such as alarms, controls, and safety systems.
PQ verifies sterilization performance with actual loads, including mapping and BI challenges.
31. What is heat distribution study?
A heat distribution study measures temperature uniformity throughout the empty autoclave chamber.
Identifies cold and hot spots using multiple calibrated sensors.
Ensures chamber delivers consistent thermal environment before load testing.
32. What is heat penetration study?
Heat penetration study evaluates how effectively steam heats the internal parts of the load.
Uses temperature sensors placed in worst-case load positions.
Confirms that load reaches and maintains sterilization temperature.
33. What are cold spots in autoclaves?
Cold spots are areas where temperature is lower than required during sterilization.
Commonly caused by trapped air, poor steam circulation, or blocked drains.
Must be identified and corrected to avoid incomplete sterilization.
34. What is the cause of cold spots?
Cold spots occur due to air pockets, improper load arrangement, wet steam, or drainage issues.
Incorrect placement of temperature probes can also indicate false cold spots.
Poor steam quality leads to uneven heating.
35. Why mapping of chamber is required?
Mapping ensures uniform temperature distribution throughout the autoclave chamber.
Mandatory for validation under GMP and regulatory guidelines.
Supports qualification of both empty chamber and loaded cycles.
36. What are biological indicators (BI)?
BI contain highly resistant bacterial spores to challenge sterilization effectiveness.
Placed at worst-case locations in the load.
Demonstrate actual microbial kill performance of the cycle.
37. What are chemical indicators (CI)?
CI are color-changing strips or labels that respond to temperature, steam, and time.
Provide quick visual confirmation of exposure to sterilization conditions.
Used for routine monitoring but not a substitute for BI.
38. What is the difference between BI and CI?
BI measure actual microbial kill using spores and confirm lethality.
CI only show exposure to sterilization parameters, not microbial death.
BI are used for validation; CI for routine checks.
39. What microorganisms are used in biological indicators?
The common organism is Geobacillus stearothermophilus.
Chosen because its spores are highly resistant to moist heat.
Ensures a strong challenge for steam sterilization cycles.
40. Why is Geobacillus stearothermophilus used in autoclave validation?
It has the highest resistance to moist heat among non-pathogenic organisms.
Provides reliable challenge to verify sterilization lethality.
Ensures that if these spores are destroyed, all other microorganisms will also be eliminated.
41. What is the minimum log reduction required for sterilization?
A minimum of 6-log reduction is required, meaning 99.9999% microbial kill.
This standard ensures destruction of highly resistant spores.
Used as the benchmark for validating sterilization cycles.
42. Why is steam quality important?
Steam quality determines the effectiveness of heat transfer.
Poor steam quality leads to wet loads, cold spots, and failed sterilization.
Ensures consistent and reliable microbial kill.
43. What are the three steam quality parameters?
Non-condensable gases, dryness fraction, and superheat.
These parameters determine whether steam can condense properly.
Directly affect the autoclave’s sterilization performance.
44. What is non-condensable gas (NCG) and why is it critical?
NCGs such as air or CO₂ do not condense and form insulating pockets.
Reduce steam penetration and temperature uniformity.
High NCG content leads to sterilization failure.
45. What is dryness fraction of steam?
Dryness fraction indicates the proportion of steam that is vapor vs moisture.
Higher dryness fraction improves heat transfer and sterilization efficiency.
Low dryness causes wet loads and temperature loss.
46. What is superheat in steam?
Superheat is the temperature above saturation temperature at a given pressure.
Superheated steam does not condense and provides poor sterilization.
Indicates steam is too dry for effective microbial kill.
47. How do you measure steam quality for autoclaves?
Steam quality test includes NCG test, dryness fraction test, and superheat test.
Performed at steam supply point before entering the autoclave.
Ensures the steam meets EN 285 or GMP requirements.
48. What is the function of temperature and pressure gauges?
They monitor real-time sterilization conditions inside the chamber.
Ensure cycle reaches required temperature and pressure.
Provide essential safety and compliance data.
49. What is the function of safety valve in autoclave?
Prevents overpressure by releasing excess steam automatically.
Protects chamber, operators, and equipment from explosion hazards.
Mandatory safety device in all pressure vessels.
50. Why is calibration required for autoclaves?
Calibration ensures sensors provide accurate readings during sterilization.
Prevents cycle failures due to incorrect temperature or pressure measurement.
Required for compliance with GMP, FDA, and internal quality standards.