Microscopy Techniques Practice Questions

Correct answer: Condenser

The condenser is a lens system located beneath the stage that collects and focuses light from the illuminator onto the specimen. Proper adjustment of the condenser is critical for achieving optimal contrast and resolution.

Correct answer: To increase resolution by reducing refraction

Immersion oil has the same refractive index as glass, which prevents light from bending as it passes from the slide to the air. This allows more light to enter the objective lens, significantly increasing the resolution of the image.

Correct answer: 450x

Total magnification is calculated by multiplying the magnification of the ocular lens by the magnification of the objective lens. In this case, 10 multiplied by 45 equals 450x.

Correct answer: Phase-contrast microscopy

Phase-contrast microscopy converts slight variations in the refractive index and thickness of different parts of a specimen into variations in light intensity. This allows for high-contrast imaging of transparent living cells without the need for lethal staining.

Correct answer: Bright specimen against a dark background

Dark-field microscopy uses a special condenser with an opaque disk that blocks direct light from entering the objective. Only light reflected or refracted by the specimen enters the lens, making the object appear bright against a black background.

Correct answer: Fluorescence microscopy

Fluorescence microscopy utilizes fluorochromes that absorb short-wavelength light (UV) and emit longer-wavelength light. When coupled with immunofluorescence (antibodies), it can pinpoint specific species or structures with high sensitivity.

Correct answer: The internal ultra-structure of thin-sectioned cells

TEM passes a beam of electrons through an ultra-thin section of a specimen. Because electrons have a much shorter wavelength than light, TEM provides extremely high resolution of internal organelles and membranes.

Correct answer: Scanning electron microscope

Scanning electron microscopy (SEM) scans the surface of a specimen coated in metal with an electron beam. The primary and secondary electrons emitted create a detailed 3D representation of the surface topography.

Correct answer: Distinguish two adjacent points as separate

Resolution (or resolving power) is the limit of usable magnification; it determines the amount of detail that can be seen. A smaller resolving power value indicates better clarity and the ability to see finer details.

Correct answer: Illuminator -> Condenser -> Specimen -> Objective -> Ocular

Light originates at the illuminator, is focused by the condenser onto the specimen, passes through the objective lens for primary magnification, and finally reaches the ocular lens for the user to view.

Correct answer: Electrons have shorter wavelengths than light

The resolving power of a microscope is inversely proportional to the wavelength of the radiation used. Since electron beams have wavelengths thousands of times shorter than visible light, they can resolve much smaller structures.

Correct answer: Confocal laser microscopy

Confocal microscopy uses a laser to scan a specimen at different depths. The pinhole aperture ensures only light from the focal plane reaches the detector, allowing for 3D reconstruction of thick samples like biofilms.

Correct answer: Prisms that split the light beam

DIC microscopy uses two beams of light split by prisms. The interference between the beams after they pass through the specimen creates an image with high contrast and a three-dimensional appearance without staining.

Correct answer: To regulate light entering the condenser

The iris diaphragm is an adjustable shutter located within or under the condenser. Opening and closing it controls the angle and amount of light, which affects both the contrast and the depth of field.

Correct answer: Scanning tunneling microscope

Scanning tunneling microscopy (STM) uses a probe that maintains a constant distance from the specimen surface by measuring electron 'tunneling' current. It provides incredibly high resolution at the atomic level.

Correct answer: The light-bending ability of a medium

Refractive index describes how much light slows down and changes direction when passing through a material. Staining specimens changes their refractive index relative to the medium, making them easier to see.

Correct answer: It decreases

As magnification increases, the area of the specimen visible (field of view) decreases. This is why it is important to center the object of interest before moving to a higher magnification.

Correct answer: The specimen must be coated with a thin layer of heavy metal like gold

Because SEM works by bouncing electrons off a surface, the specimen must be conductive. Sputter coating the sample with a heavy metal prevents charging and enhances the emission of secondary electrons.

Correct answer: The objective lens and the top of the specimen

Working distance is the space between the front lens of the objective and the cover glass or specimen when the image is in focus. This distance decreases as the magnification of the objective lens increases.

Correct answer: Bright-field compound light microscope

The bright-field microscope is the standard instrument in most labs. It shows a dark image (the stained specimen) against a bright background and is ideal for routine observations of bacterial morphology and staining characteristics.