Microscopes

Question 1

LM

Bright field

Answer 1

used for looking at stained samples

Question 2

LM

Phase contrast

Answer 2

Enhance the natural contrast of a specimen

Question 3

LM

fluorescence

Answer 3

Tag/label specific components

Light shined onto a specimen and fluorescent molecules are excited

Question 4

LM

Deconvolution

Answer 4

Remove out of focus information

Question 5

LM

Confocal

Answer 5

Remove out of focus information

Question 6

LM

what does focal length depend on?

Answer 6

curvature of a lens

A long focal length lens is less curved and a short focal length lens is highly curved

Question 7

LM

Convex lens

Answer 7

Converges parallel beams to a focal point.
A single convex lens produces an enlarged virtual image if the object is closer than the focal point.
If the object is further than the focal length a real image is formed

Question 8

LM

Concave lens

Answer 8

Diverges parallel beams

The focal point is the point they would have converged from

Question 9

LM

Objective lens

Answer 9

Lens to produce a magnified real image. A real image can only be seen if you place a screen.

Question 10

LM

Eyepiece lens

Answer 10

Produces a magnified virtual image of the real image

Question 11

Bright field microscopy

Answer 11

Light is passed through a specimen. Scattering of light and interference produces an image. This image is magnified and focused on the retina or detector

Question 12

FM

Answer 12

Fluorescence microscopy

Question 13

How does FM work?

Answer 13

Different parts of a cell are stained using specific dyes or antibodies which are attached to fluorescent molecules.
specimens are illuminated with light of a specific wavelength to excite a fluorophore. A fluorophore will emit light with a longer wavelength then the image is magnified and focused on a retina/detector.

Question 14

How are fluorescent tags attached to antigens?

Answer 14

A primary antibody will bind to an antigen. A secondary antibody with a fluorescent tag will then recognise and bind to the primary antibody.

Question 15

Filters in fluorescent microscopy

Answer 15

A light source shines onto a filter which filters out wavelengths of light that are not wanted. The specimen is illuminated with light of a specific wavelength,

Question 16

Explain fluorescence

Answer 16

A specimen is illuminated with a wavelength to excite a fluorophore/electron moving the electron into a higher energy level. As the electron moves back down an energy level the light is emitted.

Question 17

Green fluorescent protein

Answer 17

A chromophore is held within the green fluorescent protein- this is the molecule that is excited by a light wavelength.
This protein can be genetically tagged with other proteins.

Question 18

GFP and tubulin

Answer 18

Tublin can be tagged with GFP as a tubulin gene is added into GFP. Stick plasmid containing these genes into a mammalian cell, transcription and translation occurs. There is a shorter linker peptide between the tubulin and GFP genes as otherwise the tubulin would not be incorporated into microtubules.

Question 19

Confocal or deconvolution microscopy

Answer 19

Improve the clarity and resolution of fluorescence images.

Confocal microscopy pinhole prevents out of focus light reaching the detector so filters it out.

Question 20

TEM

Answer 20

Transmission electron microscopy
2D projection image of a thin specimen
Electrons scattered and shadow image created of the specimen

Question 21

SEM

Answer 21

Scanning electron microscopy

Electrons scattered across a sample surface and electrons are emitted off the surface

Question 22

TEM components compared to a light microscope

Answer 22

TEM occurs in a vacuum, no glass lenses only electromagnetic lenses

Question 23

Describe how SEM works

Answer 23

Primary beam from an electron beam is focused to a fine point
Beam scanned across a sample surface
Secondary electrons emitted from the specimen as the beam passes over it
Secondary electrons are collected by a detector and a picture is built up

Question 24

EM

condenser lens

Answer 24

Focuses electrons onto specimens

Controls diameters of beams- wide at a low magnitude

Question 25

EM

Objective lens

Answer 25

Focuses an image and magnifies it

Question 26

EM

projector lens

Answer 26

Magnification

Question 27

EM

viewing screen or photographic film

Answer 27

Viewing screen or digital camera

Question 28

Define resolution

Answer 28

The closest two objects can be and still be distinguishable. This depends on wavelength and shorter wavelengths result in better resolution.

Question 29

Resolution of a light microscope

Answer 29

200nm

Question 30

Resolution of electron microscopes

Answer 30

Electrons behave as waves so resolution = -0.005 nm but in practice this is -0.1nm