sTAINING, MOUNTING AND PREP

Question 1

Why is it important to stain cells before viewing them?

Answer 1

Staining provides contrast between different structures in the sample to make them visible so they can be identified. contrast increases as different components within a cell take up stains to different degrees.

Question 2

What is differential staining?

Answer 2

When multiple stains are used and each stains binds to a specific cell structure - staining each structure differently so structures can be easily identified.

Question 3

Acetic Orcein

Answer 3

binds to DNA

bind chromosomes a dark red

Question 4

Eosin

Answer 4

cytoplasm

dark red or p!nk

Question 5

Iodine

Answer 5

starch
blue black
violet under microscope

Question 6

iodine in potassium iodide solution

Answer 6

cellulose

yellow

Question 7

Haematoxylin

Answer 7

RNA/DNA

purple/blue

Question 8

Methylene blue

Answer 8

all purpose stain

stains DNA blue

Question 9

What is a wet mount used to observe

Answer 9

living organisms and aquatic samples

Question 10

explain process of a wet mount

Answer 10

1. use a pipette to add a drop of liquid -water or immersion oil…if stain is used it is important to ensure it is not toxic to live organism
2. use tweezers t carefully place the specimen
3. drop coverslip at a 45 degree angle to avoid air bubbles which obstruct image.

Question 11

what is a dry mount used to observe?

Answer 11

hair, pollen, parts of insects, sectioning of muscle tissue and plants

Question 12

explain the process of a dry mount

Answer 12

1. solid specimens are sectioned (cut into thin slices so light can pass them)
2. use tweezers to place specimen in the middle of the slide
3. add coverslip on top

Question 13

Explain the process of a squash slide

Answer 13

1. prepare a wet mount
2. use lens tissue to gently press down on cover slip - careful not to break it. depending on material- damage to coverslip can be avoided by squashing sample between two microscopes slides

Question 14

what would the squash slide technique be used for

Answer 14

soft samples e.g root tip squashes to look at cell division

Question 15

explain the process of a blood smear

Answer 15

edge ion a slide is used to smear the sample and crate a thin, even coating on another slide
place cover slip ontop

Question 16

example of where blood smear where be used for

Answer 16

e.g blood to view blood

Question 17

why do images of cells tend to have low contrast

Answer 17

cells do not absorb a lot of light
resolution is limited by the wavelength of light and diffraction of light as it passes sample
cytosol of cell and other structures are often transparent

Question 18

What is firstly done to prepare a sample for staining?

Answer 18

sample is first placed on slide and allowed to air dry
then this is heat-fixed by passing it though a flame. Heat fixing denatures bacterial enzymes, preventing them from digesting cell parts, which causes the cell to breakIf you heat the slide before it is completely dry, then you end up “boiling apart” the cells. The bacterial frost is air-dried before staining to fix the bacteria on the slide so that it does not wash away with a subsequent staining procedure specimen will adhere to the microscope slide and then take up stains.

Question 19

What are examples of positive dyes and how do they act?

Answer 19

direct,cationic,basic or positive dyes e.g methylene blue, basic fuchsin and crystal violet directly bind to and satin the negatively charged surface of bacterial cells.

Question 20

What are examples of negative dyes and how do they act?

Answer 20

e.g nigrosin, congo red are negatively charged and are repelled by negatively charged cytosol. These dyes stay outside of cells, leaving the cell unstained, which stand out against gained background. NEGATIve StaIn tEcNiQuE