2.1.3 Microscopy

Question 1

Define magnification

Answer 1

How many times bigger an object appears

Question 2

Define resolution

Answer 2

The ability to distinguish two points next to each other

Question 3

If the resolution of a microscopic image is too low, what can be said about the light/electrons used

Answer 3

The wavelength is too long

Question 4

Magnification equation

Answer 4

Magnification = image size / actual size

Question 5

1cm = ?mm

Answer 5

10

Question 6

1mm = ?µm

Answer 6

1000

Question 7

1µm = ?nm

Answer 7

1000

Question 8

Measurement unit conversion image

Answer 8

Question 9

How to measure an object using a microscope and graticule

Answer 9

• Line up graticule with object
• Count how many epu for the measurement
• Calibrate graticule using stage micrometer - how many µm per epu
• Multiply the number of epu by the calibration factor

Question 10

Resolution of optical microscope

Answer 10

Low

Question 11

Resolution of SEM

Answer 11

Medium

Question 12

Resolution of TEM and ADVANTAGE

Answer 12

• High
• Allows organelles to be seen in greater detail

Question 13

Magnification of optical microscope

Answer 13

Low

Question 14

Magnification of TEM

Answer 14

High

Question 15

Magnification of SEM

Answer 15

Medium

Question 16

How does a optical microscope work

Answer 16

Light ray focused onto object then onto eye using glass lenses

Question 17

How does a SEM work

Answer 17

• Electrons deflected/bounce off specimen
• Foccused by electromagnets

Question 18

How does a TEM work

Answer 18

• Electrons transmitted/pass through specimen
• Foccused by electromagnets

Question 19

Primary use of optical microscope

Answer 19

Study living cells

Question 20

Primary use of SEM

Answer 20

View 3D image

Question 21

Primary use of TEM

Answer 21

View cell ultrastructure

Question 22

Are optical microscope images in colour

Answer 22

Yes

Question 23

Are TEM images in colour

Answer 23

No

Question 24

Are SEM images in colour

Answer 24

No

Question 25

Non-limitations to optical microscopes

Answer 25

* Can be dead or alive * No vacuum * Not too thin * Stained with dye * No artefacts * Doesn't take too long

Question 26

Limitations to SEM

Answer 26

* Must be dead * In vacuum * Very thin specimin * Stained with metal * Artefacts may occur * Time consuming

Question 27

Limitations to TEM

Answer 27

* Must be dead * Must be in vacuum * Specimin must be very thin * Stained with metal * Artefacts may occur * Time consuming

Question 28

In microscope images, if the resolution is high/low what can be said about the light or electrons wavelength

Answer 28

* Low resolution = long wavelength * High resolution = short wavelength

Question 29

Describe how to extract chloroplast from a cell sample

Answer 29

* **Homogenise** cells in homogeniser * **Filter** to remove unbroken cells/debris * Place homogenate in test tube and spin in a **centrifuge** at **low** speed * Remove nuclei pellet at bottom * Respin **supernatant** in new test tube at **higher speed** * Remove chloroplast pellet

Question 30

When extracting organelles from cells during ultracentrifugation, what is the order of heaviest organelles, that must be removed

Answer 30

* Nuclei * Chloroplast - **only if plant cell** * Mitochondria

Question 31

When extracting organelles from cells, why must the solution be cold

Answer 31

Reduce enzyme activity, so organelles aren't digested

Question 32

When extracting organelles from cells, why must the solution be isotonic

Answer 32

Prevent water movement into organelles by osmosis - preventing lysis

Question 33

When extracting organelles from cells, why must the solution be pH buffered

Answer 33

* Maintain pH * Prevent denaturations of proteins and enzymes