Microscopy

Question 1

Robert Hooke

Answer 1

• used a microscope he built to examine structure of various objects including cork
• first description of cells - “these pore, or cells”

Question 2

Van Leeuwenhoek microscope

Answer 2

• built own microscope and lenses - magnify up to 500X
• described the first single celled organisms - live cells in lake water 1974
• reported “little animals” or “ animalcules”
• discovered sperm and described RBCs
• discovered bacteria in 1676

Question 3

What is Standard Light Microscopy?

Answer 3

• A broad category of microscopy that uses visible light and optical lenses to magnify specimens.
• Includes brightfield, phase contrast, DIC, etc.

Question 4

What is Brightfield Microscopy and what type of specimen is it best for?

Answer 4

• A basic light microscopy method where light passes directly through the sample.
• Best for stained or pigmented specimens due to low contrast in unstained ones.

Question 5

How does Phase Contrast Microscopy work, and what does the phase constant do?

Answer 5

• Enhances contrast by converting phase shifts from refractive index differences into brightness differences.
• The phase constant is the fixed shift introduced by the phase plate (usually 90° or 180°) to cause interference.

Question 6

What is Differential Interference Contrast (DIC) Microscopy known for?

Answer 6

Uses polarized light and interference to create sharp, high-contrast, 3D-like images of transparent, unstained specimens—ideal for live cells.

Question 7

Haematoxylin stain

Answer 7

• purple
• stains basic acidic structures eg nucleus

Question 8

Eosin stain

Answer 8

• pink
• stains basic structures eg cytoplasm/ cell walls

Question 9

Azan trichrome stain

Answer 9

• nuclei - bright red
• collagen, basement membrane, niacin - blue
• muscle, RBC - orange → red
• good for staining connective tissue and epithelium

Question 10

1878 Walther Flemming

Answer 10

• deciphered the major stages of mitosis
• used newly developed dyes such as aniline to visualise the chromosomes
• invented the terms mitosis and chromatin

Question 11

Fluorescence microscopy steps

Answer 11

1) first barrier filler: lets thru only blue light with a wavelength between 450 and 490 nm
2) bean splitting mirror: reflects lights below 510 nm but transmits light above 510 nm
3) second barrier filter: cuts out unwanted fluorescent signals, pasting the specific green fluorescein emission between 520 and 560 nm

Question 12

What is a Scanning Electron Microscope (SEM) and how does it work?

Answer 12

• A type of electron microscope that scans a specimen’s surface with a focused beam of electrons.
• It detects secondary electrons emitted from the surface to produce detailed, 3D-like images.
• Ideal for viewing surface structures at high magnification and resolution.

Question 13

Transmission electron microscope

Answer 13

• provides the highest resolution possible
• uses antibodies labelled with gold to detect cellular structures

Question 14

resolution

Answer 14

clarity of the image, minimum distance of two distinguishable points

Question 15

magnification

Answer 15

how much larger the image is compared to its real size