1. Key Area 1- Lab techniques for biologists Flashcards
define a hazard
a source of potential harm
what are the 3 main types of hazards?
substances, organisms and equipment
define a risk
the likelihood of harm arising from exposure to a hazard
what are the 3 main control measures
- appropriate handling technique
- protective clothing and equipment
- aseptic technique
what is a risk assessment
involves identifying control measures to minimise the risk
what are the 4 hazards in the lab?
- toxic or corrosive chemicals
- heat or flammable substances
- pathogenic organisms
- mechanical equipment
What are linear dilutions
they differ by an equal interval
What is a standard curve and what is it used for?
Is produced in order to determine the concentration of a solute in an unknown solution
What piece of equipment is used to determine the absorbance of a substance or its turbidity ?
a colorimeter
Define turbidity
Proportional to the density of cells in the culture
What is a buffer?
Solutions to control pH so that changes in pH don’t act as a confounding variable
What is the function of centrifugation
To separate substances of differing densities
What is the use of paper and thin layer chromatography?
To separate different substances such as: amino acids and sugars
What does the speed that each solute travels along the strip depend on? (2 reasons)
its solubility in the chromatography solvent and its differing affinity
What is affinity chromatography used for?
To separate target proteins from a mixture of proteins
How are target proteins separated in affinity chromatography?
Soluble target proteins with high affinity for the specific molecules in the solid gel column (antibodies or ligands) attach to them. Non-target proteins with weaker or no affinity are washed out, target proteins can be washed out separately producing a more pure sample.
What is the use of gel electrophoresis
Separates proteins and nucleic acids
Describe the process of gel electrophoresis?
Charged macromolecules (proteins and nucleic acids) move through an electric field applied to a buffered gel matrix (polyacrylamide gel electrophoresis or PAGE)
What separates proteins by their shape, size and charge
Native gels
Why does SDS-PAGE separate proteins by size alone?
It contains Sodium Dodecyl Sulfate (SDS) which denatures molecules passing through it and gives all molecules present a negative charge thus separating proteins only by size
Define a proteins isoelectric point (IEP)
Is the pH value at which a protein is electrically neutral with surface charges that are balanced. If the soultion is buffered to a specific pH, only the proteins that have t
Effect of IEP on proteins in solutions
If the solution is buffered to a specific pH, only the proteins that have an IEP of that pH will precipitate.
+ Proteins can also be separated using their IEPS in electrophoresis
What is the use of immunoassay techniques?
To detect and identify specific proteins
What are stocks of antibodies with the same specificity?
monoclonal antibodies
What are antibodies specific to the protein antigen linked to?
A chemical ‘label’, often a reporter enzyme that produces a colour change
When is western blotting used?
After SDS-PAGE electrophoresis
How is western blotting carried out?
Separated proteins from the gel are transferred (blotted) onto a solid medium. The protein can then be identified by using specific antibodies which have reporter enzymes attached
What is bright field microscopy used for?
To observe whole organisms, parts of organisms, thin sections of dissected tissues or individual cells
What is fluorescence microscopy used for?
Uses specific fluorescent labels to bind to and visualise certain molecules or structures within cells or tissues
What is the purpose of aseptic technique?
Eliminates unwanted microbial contaminants when culturing micro-organisms or cells
What is the equipment needed to start a microbial culture?
An inoculum of microbial cells on an agar medium or in a broth with suitable nutrients
what are the nutrients/ growth factors from required for animal cells to grow?
glucose, fatty acids, amino acids etc
Difference between primary cell lines and tumour cell lines
Primary cell lines can divide a limited number of times whilst tumour cell lines can perform unlimited divisions.
What type of dilution is often needed to achieve a suitable colony count?
serial dilution
What are haemocytometers?
Microscopic grids used to estimate cell numbers in liquid culture
What is required to identify and count viable cells?
Vital staining
