Examining Cells

Question 1

Define tisssue

Answer 1

Woven

Question 2

Name four tissue classifications

Answer 2

Muscle, nerves, epithelial, connective

Question 3

How does epithelial tissue communicate?

Answer 3

Through junctions at their lateral and basal surfaces

Question 4

Four properties of epithelial tissue

Answer 4

• on the edge of other tissue or surrounding them
• form glands when in clusters
• polarised when at surfaces
• held together by anchoring proteins

Question 5

What type of membrane does epithelial tissue have on the basal?

Answer 5

Basement

Question 6

What are connective tissues made from?

Answer 6

Cells, extra cellular proteins, glycoproteins and gel

Question 7

Name three mains cells in connective tissue

Answer 7

Fibroblasts, chondrocytes, osteocytes

Question 8

Main products of connective tissue

Answer 8

• fibres
• wax and gel materials

Question 9

What are the three muscle cells muscle tissue is made from

Answer 9

• skeletal
• cardiac
• smooth

Question 10

Three functions of muscle tissue

Answer 10

• movement
• stability
• movement of tissue
  (Made function to contract)

Question 11

Minor functions of hormones

Answer 11

• natriuretic factors (for the heart)
• myostatin (regulates the heart muscle)

Question 12

What do nerve cells congregate into?

Answer 12

Nerve fibres

Question 13

What is the main role of nerve tissue?

Answer 13

Communication

Question 14

What is the standard measurement for a cell

Answer 14

Um

Question 15

What is used to measure a cell?

Answer 15

A measurement graticule

Question 16

What is the limit of resolution

Answer 16

The smallest distance by which two objects can be separated and still be distinguishable as two separate objects

Question 17

Tissue procurement methods

Answer 17

• endometrial sample (upper uterus)
• venepuncture (intravenously)
• bone marrow aspiration

Question 18

Why do you use a buffered formalin solution during fixation?

Answer 18

To prevent the cell from swelling

Question 19

In basic terms what is fixation?

Answer 19

To preserve cells and tissue components

Question 20

What is the step after fixation?

Answer 20

Washing and dehydration

Question 21

What is the most common fixation chemical and why is it isotonic with the intracellular fluid?

Answer 21

Formalin
Allows better penetration of the formaldehyde

Question 22

Name two properties of the sample during fixation and why

Answer 22

Small - fixation needs to be rapid
24-48 hours - no longer or they will shrink

Question 23

X

Question 24

Why use paraffin wax?

Answer 24

It is fluid when heated but hardens when solid

Question 25

What happens in washing?

Answer 25

The tissue is washed in a series of alcohol solutions Then in solvents

Question 26

What is the key characteristic of the solvents

Answer 26

Miscible with both alcohol and hot paraffin wax (completely dissolved in each other)

Question 27

What happens to tissue after being washed with the solvent?

Answer 27

It is immersed in hot paraffin wax overnight Then placed in a mold More hot wax over to completely cover Set aside to harden fully (can now be stored indefinitely)

Question 28

What is used to cut the hardened specimen?

Answer 28

Microtome

Question 29

What blade can a microtome use?

Answer 29

Steel Glass Diamond knife

Question 30

Where is the cut specimen then put and why?

Answer 30

A warm water bath (using a fine paintbrush) The surface tension causes the section to stretch slightly - cutting artefacts are reduced

Question 31

What can you cover the microscope slid with and why?

Answer 31

A sticky substance Specimen adheres when dry

Question 32

What must you do to the paraffin section before you can look at it through a microscope?

Answer 32

Dissolve Rehydrate With different percentage of alcohols Stain it

Question 33

What are common stains used?

Answer 33

Haematoxylin Eosin

Question 34

How do you stain using haematoxylin? And what does it bind to?

Answer 34

Immerse in an aqueous solution of haematoxylin It is basic, so will bind to acidic structures (DNA, RNA)

Question 35

How do you then stain with eosin? And what does it bind to?

Answer 35

Wash it in water and alcohol - eosin is more soluble in alcohol Immerse in eosin Binds to basic structures because is acidic (cytoplasm, collagen - intra and extra cellular proteins) Wash with more alcohol because mounting

Question 36

How do you mount a specimen?

Answer 36

On a non-aqueous mounting medium Apply a cover slip Wait for the solvents to evaporate

Question 37

What do immunohistochemistry and immunofluorescence use?

Answer 37

Labelled antibodies Targeting specific antigens

Question 38

What are the antibodies in immunofluorescence bound to?

Answer 38

Dyes

Question 39

What can you add to the after adding a primary antibody? What is this called?

Answer 39

Enzyzme activated secondary antibody - precipitate a coloured product at the site of interaction Indirect immunohistochemistry (even more sensitive)

Question 40

Why are antigen retrieval methods used?

Answer 40

Protein structure may have been altered by tissue processing procedures

Question 41

Describe antigen retrieval

Answer 41

Heated - partial digestion In the presence of a weak acid

Question 42

Why does antigen retrieval work on less robust tissue samples? So what would you use?

Answer 42

They cannot be heated A frozen section

Question 43

Why can more components be seen using an electron microscope?

Answer 43

Higher resolution

Question 44

What is the limit resolution mean?

Answer 44

Minimum distance at which two objects can be distinguished

Question 45

Five comparisons of light and electron microscopes

Answer 45

L - colour E - black and white L - cheaper and easier preparation L - living, moving L - lower magnification L - lower resolution

Question 46

What type of microscope is confocal?

Answer 46

Scanning

Question 47

How does a confocal microscope work?

Answer 47

1. Laser excites dye - electrons raised to higher energy level 2. Electrons relax back to ground state 3. Light with a higher wavelength emitted 4. Emitted light sent to mirrors and a pinhole screens to CMOS detector

Question 48

Why can a confocal microscope give a 3D image?

Answer 48

Multiple images taken Computer places them in order Creating a Z stack

Question 49

What are confocal microscopes used for?

Answer 49

Evaluating eye diseases

Question 50

What are the five steps on preparing live cells?

Answer 50

1. Cutting and dicing 2. Collagenase and DNAse (enzymes used to ‘break’ the cells apart) 3. Centrifugation 4. Put in appropriate growth medium 5. Culture cells

Question 51

What sort of microscope is used to look at live cells?

Answer 51

Phase contrast

Question 52

Three advantages of cell culture

Answer 52

- control over physical environment - homogeneity (one type) - reduced need for animals

Question 53

Three disadvantages of cell cultures

Answer 53

- hard to maintain (sterile) - small amount at a high cost - dedifferentiation

Question 54

What is dark field?

Answer 54

Illuminate sample of live cells Light not collected by objective lens - black background with bright objects

Question 55

What type of electron microscope gives a 3D image?

Answer 55

Scanning

Question 56

Why can you see an oocyte?

Answer 56

Bigger than the resolving power of the eye

Question 57

What is H&E?

Answer 57

Haematoxylin Eosin

Question 58

Smallest organelle to see with a light microscope?

Answer 58

Nucleolus

Question 59

Why is hard to look at fat under the microscope?

Answer 59

Fat dissolves in the alcohol during the preparation method When using H&E

Question 60

What is an erythrocyte?

Answer 60

Red blood cell