BACE Microscopy and Imaging Techniques 3 — Questions and Answers
Question 1: What is a 'wet mount' preparation in light microscopy?
- A specimen suspended in a drop of liquid between a slide and coverslip (Correct answer)
- A slide that has been washed with water before use
- A preparation using water-soluble stains
- A slide stored in humid conditions
Correct answer: A specimen suspended in a drop of liquid between a slide and coverslip
A wet mount places a specimen in a drop of water or buffer covered by a coverslip, allowing observation of live or unstained specimens.
Question 2: What does SEM (Scanning Electron Microscopy) primarily image?
- The surface topography of specimens (Correct answer)
- The internal structure of cells
- Fluorescently labeled proteins
- DNA and RNA molecules directly
Correct answer: The surface topography of specimens
SEM scans a focused electron beam over the specimen surface, detecting secondary electrons to create three-dimensional surface images.
Question 3: Which objective lens would provide the greatest working distance from the lens to the specimen?
- 4x (scanning objective) (Correct answer)
- 10x (low power)
- 40x (high dry)
- 100x (oil immersion)
Correct answer: 4x (scanning objective)
Lower magnification objectives like the 4x scanning objective have the greatest working distance, while higher magnification objectives are much closer to the specimen.
Question 4: In fluorescence microscopy, what does a fluorophore do when excited by light of the correct wavelength?
- It absorbs the excitation light and emits light at a longer wavelength (Correct answer)
- It absorbs all wavelengths of light equally
- It reflects the excitation light back to the detector
- It generates heat to warm the specimen
Correct answer: It absorbs the excitation light and emits light at a longer wavelength
A fluorophore absorbs photons at its excitation wavelength and re-emits photons at a longer (lower energy) emission wavelength, a process called fluorescence.
Question 5: What is the correct order of steps when beginning microscopy observation of a new slide?
- Place slide → use lowest power objective → focus with coarse adjustment → switch to higher power using fine adjustment only (Correct answer)
- Place slide → use highest power objective → focus with fine adjustment → switch to lower power
- Focus without slide → place slide → use highest power immediately
- Place slide → focus with oil immersion objective first → then use dry objectives
Correct answer: Place slide → use lowest power objective → focus with coarse adjustment → switch to higher power using fine adjustment only
Always start with the lowest power objective to locate the specimen and achieve initial focus before progressing to higher magnification objectives.
Question 6: Which type of microscopy is best suited for creating three-dimensional reconstructions of thick fluorescent specimens?
- Confocal laser scanning microscopy (Correct answer)
- Bright-field microscopy
- Dark-field microscopy
- Simple light microscopy
Correct answer: Confocal laser scanning microscopy
Confocal microscopy acquires serial optical sections through the specimen that can be digitally stacked to reconstruct three-dimensional structures.
BACE Microscopy and Imaging Techniques 3
What is a 'wet mount' preparation in light microscopy?