BACE Aseptic Technique and Contamination 2 — Questions and Answers
Question 1: What is the primary purpose of a biosafety cabinet (BSC) in a biotechnology laboratory?
- To store hazardous chemicals safely
- To provide a sterile work environment and protect personnel from biohazards (Correct answer)
- To incubate cell cultures at controlled temperatures
- To centrifuge biological samples at high speed
Correct answer: To provide a sterile work environment and protect personnel from biohazards
A biosafety cabinet provides a sterile, HEPA-filtered work area that protects both the samples and the operator from contamination and biohazardous aerosols.
Biosafety cabinets use HEPA filtration and laminar airflow to create a clean work zone. Class II BSCs protect both the product (from environmental contamination) and the worker (from biohazardous aerosols), making them the standard for cell culture and microbiology work in biotech labs.
Question 2: Which of the following actions would most likely introduce contamination into a sterile cell culture flask?
- Working inside a laminar flow hood
- Briefly resting the culture flask cap on a clean bench surface outside the hood (Correct answer)
- Flaming the neck of a glass flask before pouring
- Using sterile disposable pipettes
Correct answer: Briefly resting the culture flask cap on a clean bench surface outside the hood
Placing a culture flask cap on an external bench surface exposes it to environmental microorganisms, which can then contaminate the culture when the cap is replaced.
Cap-down contamination is one of the most common sources of culture contamination. Any sterile surface that contacts a non-sterile environment picks up microorganisms. Best practice is to hold caps in your hand or keep them inside the BSC at all times while working with open vessels.
Question 3: What does the term 'sterile technique' specifically mean in a laboratory context?
- Wearing gloves and a lab coat
- Preventing any microorganisms from entering a sterile environment or culture (Correct answer)
- Cleaning benches with 70% ethanol before work
- Autoclaving all solutions used in experiments
Correct answer: Preventing any microorganisms from entering a sterile environment or culture
Sterile technique refers to the set of practices used to prevent any microorganisms from contaminating sterile materials, media, cultures, or equipment.
Sterile technique encompasses all practices that prevent microbial contamination: working in certified clean environments, using sterile materials, minimizing open-vessel exposure time, and proper handling of caps, pipettes, and equipment. It goes beyond PPE to encompass work habits and environmental controls.
Question 4: How often should the interior surfaces of a biological safety cabinet be decontaminated with 70% ethanol?
- Only when visibly soiled
- Once per week regardless of use
- Before and after each use session (Correct answer)
- Once per month during scheduled maintenance
Correct answer: Before and after each use session
BSC interior surfaces should be wiped with 70% ethanol before starting work and again after completing work to ensure the work surface remains free of contamination.
Pre- and post-use decontamination is standard practice for BSCs. Before use, it removes any particles or organisms that settled between sessions. After use, it eliminates any biological material that could pose a risk to the next user. 70% ethanol is preferred over 100% because the water content aids protein denaturation.
Question 5: A laboratory technician notices that a cell culture flask contains turbid (cloudy) media that was previously clear. What is the most likely explanation?
- The cells are growing normally and have reached confluence
- Microbial contamination has occurred in the culture (Correct answer)
- The media pH has shifted due to CO2 levels
- The cells have differentiated into a new cell type
Correct answer: Microbial contamination has occurred in the culture
Sudden turbidity in previously clear culture media is a classic sign of microbial (bacterial or yeast) contamination, which rapidly multiplies and clouds the media.
Turbid media indicates a very high microbial load — typically bacteria or yeast that have contaminated the culture. Unlike mammalian cells, which grow slowly and adhere to surfaces, bacteria and yeast grow rapidly in suspension and can cloud media within 24–48 hours. Contaminated cultures must be discarded and autoclave-decontaminated before disposal.
Question 6: What is the correct order of operations when beginning work in a biological safety cabinet?
- Load materials then turn on blower then wait 5 min then begin work
- Turn on blower then wait 5 min for airflow to stabilize then wipe with 70% ethanol then begin work (Correct answer)
- Wipe with ethanol then turn on blower then immediately begin work
- Turn on UV light then turn on blower then begin work immediately
Correct answer: Turn on blower then wait 5 min for airflow to stabilize then wipe with 70% ethanol then begin work
The BSC blower must be running and airflow must stabilize before work begins; surfaces are then decontaminated with 70% ethanol before introducing materials.
Proper BSC startup requires: (1) turn on blower and allow 5-10 minutes for HEPA-filtered laminar airflow to stabilize, (2) wipe all interior surfaces with 70% ethanol and allow to dry, (3) load only required materials, then (4) begin work. UV lights are used between sessions, not during active work, and are not a substitute for chemical decontamination.
BACE Aseptic Technique and Contamination 2
What is the primary purpose of a biosafety cabinet (BSC) in a biotechnology laboratory?