Microscopy Techniques Flashcards
7 cards from real Histology Certification practice questions. Tap to flip, then mark Knew It or Still Learning — missed cards come back until you master them.
Read the first 7 Microscopy Techniques flashcards as text
In scanning transmission electron microscopy (STEM), what detector is used for high-angle annular dark-field (HAADF) imaging?
Answer: High-angle annular ring detector that collects scattered electrons
HAADF-STEM uses an annular detector positioned at high angles to collect Rutherford-scattered electrons, producing Z-contrast images where heavier elements appear brighter.
What is the critical point drying method used for in specimen preparation for SEM?
Answer: To remove water without surface tension damage by transitioning through the critical point of CO2
Critical point drying removes liquid from specimens by taking CO2 above its critical point (31°C, 73 atm), eliminating the liquid-gas interface and preventing collapse of delicate structures.
In atomic force microscopy (AFM), which imaging mode minimizes sample damage by briefly tapping the surface rather than dragging across it?
Answer: Tapping (intermittent contact) mode
Tapping mode AFM oscillates the cantilever near its resonant frequency, briefly contacting the surface at the bottom of each oscillation, greatly reducing lateral frictional forces that could damage soft biological samples.
What is the purpose of sputter coating specimens before SEM examination?
Answer: To apply a thin conductive metal layer to prevent charge buildup
Sputter coating deposits a thin layer of conductive metal (gold, platinum, or palladium) on non-conductive biological specimens, preventing charging artifacts caused by electron beam accumulation.
Which fluorescence microscopy technique uses total internal reflection to selectively illuminate only fluorophores within ~100 nm of the cover glass surface?
Answer: Total internal reflection fluorescence (TIRF) microscopy
TIRF microscopy uses an evanescent wave created at the glass-water interface under total internal reflection conditions to excite only fluorophores extremely close to the surface, drastically reducing background fluorescence.
In two-photon excitation fluorescence microscopy, what is the primary reason for reduced photobleaching outside the focal plane?
Answer: Significant two-photon absorption probability exists only at the focal point where photon density is highest
Two-photon excitation requires near-simultaneous absorption of two photons, which occurs with appreciable probability only at the focal spot where photon flux is extremely high, confining excitation and photobleaching to that point.
What is the working distance of a microscope objective?
Answer: The free distance between the front lens element and the focal plane of the specimen
Working distance is the gap between the front lens of the objective and the coverslip surface when the specimen is in focus; it decreases as magnification and NA increase.