Practical Exam Flashcards
P1: Method of semen collection for boars
“Gloved hand method”
P1: Important points when collecting semen (3)
- Avoid contamination of semen (clean the prepuce of the male).
- Use clean, sterile, dry and warm semen collection vessels; avoid cold shock.
- Ensure dummy sow is secure and non-slip mat is in place
P1: Why should you work in groups of three when inseminating?
One inseminates
One applies back pressure
The other assists.
P2: What temperature should ram ejaculate be kept at in water bath?
30 degrees
P2: What are two ways of estimating concentration of semen?
- Haemocytometer
2. Photocolorimeter
P2: Types of abnormalities in sperm:
- tailless (decapitated)
- coiled tail
- tapering head
- broken at neck or midpiece
- small or large head
- enlarged midpiece
- double head
- double tail
- protoplasmic droplet
- abnormal arosome
P2: Wave motion
5 - Dense, very rapid vigorous waves, changing direction very rapidly, individual spermatozoa cannot be seen; 80% or more of the spermatozoa are alive and active.
4 - Waves and eddies are not so rapid as for score 5; about 65 - 80% of spermatozoa are active.
3 - 45 - 65% of spermatozoa are active. Only a few small and slow waves, or no waves. Individual spermatozoa can be observed.
2 - 20 - 45% of spermatozoa are alive, motility is poor. No waves are formed.
1 - Very few sperm cells show signs of life with very weak and non- progressive movement.
0 - No motility. All spermatozoa are dead.
P2: Motility of spermatozoa (Types of motion):
a. Progressive forward movement;
b. Rotary movement: individual sperm move in a circle;
c. Oscillatory, or convulsive movement, without progress and change in position.
P2: When counting sperm heads using haemocytometer, how do you increase uniformity?
- Heads on top and right lights counted IN
- Heads on bottom and left lines counted OUT
P2: When counting sperm heads using haemocytometer,, how many squares are counted?
- 4 corners and a middle square.
P2: What is the volume of 25 large squares on haemocytometer?
1/10,000 ml
P2: What is the calculation to determine the number of sperm/ml in original sample from haemocytometer counts?
No. sperm/ml = No. sperm in (5 squares x 5) x dilution rate (200) x 1/volume of 25 squares (10,000)
P3: Cryobiology
The study of the effect of sub zero temps on biological materials
P3: Cryopreservation
The storage of biological materials at sub zero temps
P3: Why cryopreserve
Increases the longevity of a sample
- preservation of tissue or cells e.g. sperm, oocytes, embryos
P3: Role of cryoprotectant
Protects cells against freezing injury
- Decrease freezing point of a solution
- Maintain membrane fluidity
P3: To allow cells to remain viable throughout freezing and thawing, it is important to:
- Minimise damage due to dehydration
- Minimise damahe due to ice formation
P3: What effect does contamination of water (e.g. salt addition) have on it’s freezing/thawing point?
It DECREASES the freezing/thawing point
P3: Define osmolarity
The concentration of a solution, often defined as osmols/L
P3: Define tonicity
The osmotic pressure of a solution, often expressed as relative to an intercellular environment (e.g. hypo, iso, hypertonic)
P3: Define pH buffer
A solution which reduces the change of pH upon addition of small amounts of acid or base.
P3: Impact of freezing too slowly
= Dehydration
- Extracellular ice formation
- Slow increase in concentration of extracellular solution
- Water rushes out of cell
P3: Impact of freezing too fast
= Explosion
- Intracellular ice formation
- Intracellular ice expansion
- Physical damage and cell death
P3: What does the optimal freezing rate depend on?
- Cell size (larger cells like embryos have a slower cooling rate, smaller cells like sperm have a faster cooling rate)
- Extracellular solution (presence of cryoprotectant)
P3: Role of egg yolk in cryopreservation
- Active ingredient = low density lipoproteins (LDL’s)
- LDL’s stabilise the structure and prevent loss of lipids from membrane during freezing
P3: Vitrification
Freezing in the absence of ice
OR solidification of liquid without crystallisation
P3: Pros and cons of vitrification
For - Improved survival rates - Application to whole tissue not just cells Against - Technically difficult - Questionable biosecurity
P3: Seeding
= Facilitated ice formation or ice nucleation
Deliberate initiation of ice formation at predefined temperatures allows for more controllable and reproducible freezing conditions
P3: DMSO and Glycerol as cryoprotectants
- Penetrating cryoprotectants
- DMSO: Permeates rapidly, embryo freezing
- Glycerol: Permeates slowly, sperm freezing
P3: Sucrose and egg yolk as cryoprotectants
- Non-penetrating cryoprotectants
- Sucrose: Enhances effect of penetrating cryoprotectant
- Egg yolk: Improves membrane fluidity
