Test 2 - Microscopy

Question 1

Why does an image look different in bright-field, phase - contrast, and differential - interference contrast microscopy?

Answer 1

Bright field microscopy has no interference just a series of lens. Phase - contrast uses a phase plate in the objective, and an annular diaphragm that shifts the light to give better definition. Differential interference - contrast uses light that is split into two perpendicular components before passing them through the specimen. Gives a 3-D look that allows for the examination of thick and detailed structures thanks to contrast.

Question 2

Describe how GFP works?

Answer 2

• When hit with UV light it fluoresces green
  
  • Can be used to tag proteins that don’t have antibodies.

Question 3

Bright-field microscopy

Answer 3

Basic image, very little detail or definition

Question 4

Phase - contrast

Answer 4

Cone of light illuminates specimen

Question 5

Differential - interference

Answer 5

Allows you to see alot more detail, including: Small details and thick objects. Works by having differential height.

Question 6

Conventional fluorescence microscopy

Answer 6

Deeper imaged volume that dilutes the picture with background noise.

Question 7

Confocal fluorescence microscopy

Answer 7

• It only obtains images from a specific focal plane and excludes light from other focal planes.
  
  • Allows you to see where a protein is in a cell.
• Collect series of images vertically
  
  • Can be used to generate an accurate 3-D representation of specimen.

Question 8

Two Photon Excitation Microscopy

Answer 8

• Two photons of lesser energy can excite an electron the same as one photon.
• Benefits:
  
  • Visualize thicker specimens
  • Can be used on living organisms.
  • Can be focused onto a single plane

Question 9

Describe how FRET works?

Answer 9

• Fluorescence energy resonance transfer​
  
  • Used to determine if two proteins interact in a cell.
    
    • Excitation of the primary protein causes the primary protein to emit a wavelength that will excite the secondary protein.
    • If the secondary protein light is seen it means that the proteins are extremely close to one another.
• Uses GFP

Question 10

What is FRAP?

Answer 10

• Fluorescent recovery after photobleaching
  
  • If recovery of fluorescence occurs it means that things are moving in the cell membrane.

Question 11

Transmission Electron Microscopy

Answer 11

Allows you to make a platinum cast of specimen and you look at the cast directly.

Question 12

Scanning Electron Microscopy

Answer 12

Much higher detail than a TEM.

Looking through a Electron detector.