BL10 L2 Examining Cells And Microscopy

Question 1

List the 4 types of tissues

Answer 1

• striated muscle
• nerves
• connective
• epithelial

Question 2

Resolution definition

Answer 2

Smallest distance that two objects can be separated and seen as two different objects

Question 3

Mili

Answer 3

1mm = 1x10^-3m

Question 4

Micro

Answer 4

1um = 1x10^-6m

Question 5

Nano

Answer 5

1nm = 1x10^-9

Question 6

List 5 biopsy techniques

Answer 6

• surgery and later dissections
• scraping methods e.g. scalpel, scrapes, curettes
• sharp needles e.g. needle/ punch biopsy, pipelle
• direct venepuncture e.g for blood smear
• tranvascular: device travels through BVs to site of biopsy e.g. liver, lungs, brain, heart

Question 7

Examples of scaring biopsy methods

Answer 7

Curettes
Scalpel scrapes

Question 8

Examples of sharp needle biopsy techniques

Answer 8

Needle biopsy
Punch biopsy
Pipelle

Question 9

What can a pipelle be used for?

Answer 9

Endometrial biopsy sample
Endometrium-Uterus lining

Question 10

What does transvascular mean and what can it be used to take a biopsy of?

Answer 10

Device travels through BVs to biopsy site
Kidney, lungs, brain, heart

Question 11

2 reasons for fixation

Answer 11

• samples need to be translucent + thin
• protect against microbes

Question 12

How thin do samples need to be for fixation

Answer 12

2-10 micrometres

Question 13

Why do samples need to be translucent and thin?

Answer 13

To avoid diffraction of light and prevent blurry image

Question 14

Why do samples need to be fixated to protect against microbes?

Answer 14

Once of out the body, samples are no longer under the protection of the immune system

Question 15

What fixative is used?

Answer 15

Formalin solution - 37% solution of formaldehyde

Question 16

How long do you fixate for?

Answer 16

24-48 hours only

Question 17

What happens if you fixate for too long?

Answer 17

Shrinkage occurs due to dehydration
Causes fixation artefacts

Question 18

What causes fixation artefacts?

Answer 18

Fixation for longer than 48 hours
Causes shrinkage due to dehydration

Question 19

How does paraffin wax embedding occurs?

Answer 19

• dehydrating in varying alcohol cones
• immersed in hot paraffin wax overnight
• tissue orientated in mould
• add more wax
• allow to solidify
• slice thinly using microtome

Question 20

What machine is used to slice samples thinly?

Answer 20

Microtome

Question 21

Frozen section process

Answer 21

• specimen frozen rapidly
• frozen specimen cut with microtone
• stained with H+E

Question 22

Difference between paraffin embedded tissue section and frozen sample

Answer 22

Paraffin - fixed tissue, 24-48 hours to make, permanent, clear/detailed
Frozen sample - fresh tissue (preserves lipid containing organelle), 10-20 mins to makes, lasts a few months, lacks detail

Question 23

What colour does haematoxylin stain and what does it stain?

Answer 23

Blue
Nucleus - binds to acidic things e.g DNA, RNA

Question 24

What colour does eosin stain and what does it stain?

Answer 24

Pink
Cytoplasm, extracllular matrix

Question 25

What colour does H+E stain and what difference does it make to only using one?

Answer 25

Purple Nuclei stained clearer Cytoplasm more detailed

Question 26

Tissue prep for light microscope

Answer 26

- fix with formalin - embed with paraffin wax - stain e.g. H&E, methylene blue

Question 27

Tissue preparation for electron microscope

Answer 27

- fix with glutaraldehyde - embed with epoxy resin - stain e.g. osmium tetroxide

Question 28

What has a high resolution, light or electron microscope?

Answer 28

ELECTRO MICROSCOPE Light- 0.25 MICROmetres Electron- 0.25 NANOmetres

Question 29

What has a higher magnification, light or electron microscope?

Answer 29

ELECTRON MICROSCOPE Light- x600 Electron- x500,000

Question 30

What can be viewed with light microscopes compared to electron microscopes?

Answer 30

Light- living and moving Electron- dead and inert (have to be vacuumed)

Question 31

Colours seen in light vs electron microscope

Answer 31

Light- natural colour Electron- black and white

Question 32

Financial costs of light vs electron microscopy

Answer 32

Light- cheap and easy prep Electron- expensive and difficult prep

Question 33

What type of image does SEM give?

Answer 33

3D Black and white

Question 34

What image does TEM give?

Answer 34

2D Black and white

Question 35

Advantage of cell culture

Answer 35

- complete control over physical environment - homogeneity (the same) of sample - reduced need for animal models

Question 36

Disadvantages of cell culture

Answer 36

- hard to maintain so skilled worker needed - high cost for small amount - 3D is lost - aneuploidy - abnormal number of chromosome in haploid organism

Question 37

What can be visualised in dark field and how does it work?

Answer 37

- living cells - illuminates sample with light that won’t be collected by objective lens so wont form part of image - very dark background with bright objects on

Question 38

How does confocal microscopy work?

Answer 38

- laser excites fluorescent dye - electrons raised to higher energy level - wavelength is emitted as electron returns to ground state - emitted light is sent through mirrors and pinhole screen to CMOS detector

Question 39

What can confocal microscopy be used for clinically?

Answer 39

Evaluation of various eye diseases