Laboratory Techniques For Biologists

Question 1

What can present a hazard?

Answer 1

Substances, organisms and equipment in a laboratory

Question 2

What is a Hazard?

Answer 2

• A toxic or corrosive chemical
• Heat or flammable substances
• Pathogenic organisms
• Mechanical equipment

Question 3

What is a risk?

Answer 3

The likelihood of harm arising from exposure to a hazard

Question 4

How can Biologists identify and control measures to minimise risks and reduce hazards?

Answer 4

Creating a risk assessment

Question 5

What is a control measure?

Answer 5

• Using appropriate handling techniques
• Protective clothing and equipment
• Aseptic techniques

Question 6

What are linear dilutions?

Answer 6

When dilutions differ by an equal amount e.g. 0.1, 0.2, 0.3

Question 7

What are log/serial dilutions?

Answer 7

Dilutions that differ by a constant proportion e.g. 0.1, 0.01, 0.001

Question 8

What is a standard curve?

Answer 8

Plotting known measurements onto a graph to determine unknown measuremeants

Question 9

How does a buffer control pH?

Answer 9

A buffer is a solution where adding acids or alkalis has very little effect on the pH. This allows pH in a reaction mixture to be kept constant.

Question 10

What can a colorimeter be used for?

Answer 10

A colorimeter can be used to quantify concentration and turbidity (the cloudiness)

Question 11

How do colorimeters work?

Answer 11

• Before use the colorimeters need to be calibrated with an appropriate blank sample to provide a baseline reading.
• Light is split into its component colours and filtered so there is one wavelength of light.
• This is then passed through the sample solution where a detector picks up how much light has been absorbed by the sample or transmitted (passed through)

Question 12

What can absorbance be used for?

Answer 12

To determine the concentration of a coloured solution using a suitable wavelength filter

Question 13

What can percentage transmission be used for?

Answer 13

Percentage transmission can be used to determine turbidity, such as cells in suspension

Question 14

How does centrifuge separate substances?

Answer 14

Samples are spun at incredibly fast speeds. More dense components settle to form the pellet whilst less dense components remain in the supernatant.

Question 15

How does paper and thin layer chromatography separate different amino acids and sugars?

Answer 15

The speed that each solute travels along the chromatogram depends on its differing solubility in the solvent used.

Question 16

How does affinity chromatography separate proteins?

Answer 16

A solid matrix or gel is created with specific molecules, usually receptors, bound to it. Soluble, target proteins in a mixture, with a high affinity for these molecules, become attached to them as the mixture passes down the column. Other non-target molecules with a weaker affinity are washed out.

Question 17

How does Gel electrophoresis separate proteins and nucleic acids?

Answer 17

Samples are loaded into wells in a gel which will have an electric current running through it. As they are usually charged molecules they will move towards the opposing charge. Smaller molecules will travel faster than larger molecules so they will travel further.

Question 18

How do native gels separate proteins?

Answer 18

They separate proteins by their shape, size, and charge, They do this by ensuring they do not denature the molecule.

Question 19

How do SDS-PAGEs separate proteins?

Answer 19

They separate by size alone. They do this by giving all molecules an equally negative charge and denature them.

Question 20

How do IEPs separate proteins from a mixture?

Answer 20

Proteins with different IEPs are loaded into a gel matrix. The proteins migrate towards the charges until they reach an area with the pH or their IEP. Proteins stop migrating as they have no net charge and precipitate out.

Question 21

What happens if the solution is buffered (IEP)?

Answer 21

Only the proteins that have an IEP of that pH will precipitate out.

Question 22

Explain the relationship between electrophoresis and IEPs

Answer 22

Proteins can be separated using their IEPs in electrophoresis

Question 23

What is immunoassay?

Answer 23

Its a technique which is used to detect and identify specific proteins.

Question 24

What do immunoassay techniques use?

Answer 24

Stocks of antibodies with the same specificity, known as monoclonal antibodies.

Question 25

What is linked to a chemical label?

Answer 25

An antigen that is specific to an antibody.

Question 26

What is a chemical label?

Answer 26

Often a reporter enzyme producing a colour change,.

Question 27

What is immunoassay used for?

Answer 27

Using a specific antigen to detect the presence of antibodies to test for diseases.

Question 28

What is western blotting?

Answer 28

A technique used after SDS-PAGE electrophoresis.

Question 29

How does western blotting work?

Answer 29

Once the proteins have been separated in the gel they are transferred or blotted onto a solid medium. The proteins can then be identified using specific antibodies with reporter enzymes attached.

Question 30

What is bright field microscopy?

Answer 30

Form of microscopy commonly used to observe whole organisms, parts of organisms, thin sections of dissected tissue or individual cells.

Question 31

What is Fluorescence Microscopy?

Answer 31

Form of microscopy which uses specific fluorescent labels to bind to and visualise certain molecules or structures within cells or tissues.

Question 32

What does an aseptic technique do?

Answer 32

Eliminates unwanted microbial contaminants when culturing micro-organisms or cells

Question 33

How can a microbial culture be started?

Answer 33

Using an inoculum of microbial cells on an agar medium, or in a broth with suitable nutrients.

Question 34

What are animal cells grown in?

Answer 34

A medium containing growth factors from serum. The growth factors promote growth and proliferation.

Question 35

What is the difference between primary and tumour cells

Answer 35

Primary cell lines can divide a limited number of times, whereas tumour cell lines can perform unlimited divisions.

Question 36

How can the number of colony-forming units be counted and the density of the cells in the culture be estimated?

Answer 36

Plating out of a liquid microbial culture on solid media.

Question 37

Why is a serial dilution often used?

Answer 37

To achieve a suitable colony count

Question 38

What is a haemocytometer and what does it do?

Answer 38

Specialised microscope cells which are used to estimate cell numbers in a liquid culture.

Question 39

What is needed in order to identify and count living cells?

Answer 39

Vital staining is required, any living cells will absorb the stain whilst non-living cells do not.