Workshop 1- Centrifugation

Question 1

What is a centrifuge?

Answer 1

A machine that uses a force to pull dense objects away from the center to separate the particles

Question 2

Label parts do Centrifuge

Answer 2

Cover or Lid
Body (control temp, speed, time)
Chamber for rotor (where centrifuge is placed)
Spindle

Question 3

What would happen if the centrifuge is not balanced?

Answer 3

The spindle will break, the whole centrifuge May get damaged, personal injury.

Question 4

Label parts of tube

Answer 4

Liquid= centrifugate/supernatant

Solid=Pellet

Question 5

What determines sedimentation rate?

Answer 5

size of particle
Shape of particle
Density of particle
Density of liquid
Viscosity of liquid

Gravitational force (Xg)

Question 6

What is the formula to calculate the velocity of sedimentation?

Answer 6

V= d ²(p(particle)-p(liquid)*g
———————————
18 μ

d=diameter of particle
P=density
G= gravitational force
μ= viscosity of liquid

Question 7

Describe TableTop Clinical Centrifuge

Answer 7

Can run at a speed of up to 3000 rpm
Can hold tubes of 15-50ml
Can only pellet cells

Question 8

Describe Tabletop Microfuges

Answer 8

Can run up to 14,000rpm

Ideal for Pelleting biomolecules (DNA,RNA, proteins), and virus isolation

Question 9

Describe TableTop Ultracentrifuge

Answer 9

Can rip up to speed of 150,000RPM
Allow FRACTIONATION of biomolecules eg;
Plasmid DNA
Chromosomal DNA
RNA

Question 10

Describe floor model high speed centrifuge

Answer 10

Can run up to 30,000RPM = 100,000xg

Suffice to to pellet Cell Nuclei & Biomolecules

Question 11

Describe floor model High capacity Centrifuges

Answer 11

Can run up to 5,000rpm (7,480xg)
Has a maximum capacity of 9LITERS
Allows processing of 36L/hour when harvesting bacteria

Question 12

Describe floor model Uktrcentrifuge

Answer 12

Can run up to 130,000 RPM
Allow FRACTIONATION of Biomolecules eg ;
Plasmid DNA
Chromosomal DNA
RNA

Question 13

Order of organelles according to SIZE

Answer 13

Plasma Membrane sheets (3-20 μm)
Nucleus (4-12 μm)
Golgi Tubules (1-2 μm)
Mitochondria (.4-2.5 μm)
Lysosome (.4-.8 μm)
Vesicles (.05-.3 μm)

Question 14

How to prepare sample for cell fractionation

Answer 14

Homogenize the tissue/cells in a blender to break them up

Centrifuged at different speeds and durations

Question 15

How to pellet

• Nuclei & Cell debris
• Mitochondria and chloroplasts
• Microsomes (pieces of plasma membrane and cell’s internal membranes)
• Ribosomes

Answer 15

Nuclei (1000g) for 10 min
Mitochondria (20,000g) for 20 min
Microsomes (80,000g) for 60 min
Ribosomes (150,000g) for 3 hours