Unit 1- Laboratory techniques for biologists Flashcards
What is a hazard?
A hazard is anything that poses a potential threat to individuals or environment.
What is a risk?
Risk is the likelihood of harm arising from exposure to a hazard.
What reduces the risk of a hazard?
A risk assessment
What is a risk assessment?
Involves identifying control measures to minimise risk.
What are examples of control measures?
- Using appropriate handling techniques
- Protective clothing and equipment
- aseptic technique
What is linear dilution?
A series that differs by equal intervals for example, 0.1 0.2 0.3 etc
What is a log/serial dilution?
A series that differs by a constant proportion for example 10^-1, 10^-2, 10^-3
What is a standard curve?
Involves plotting measured values for known concentrations to produce a line or curve.
What does a standard curve allow you to do?
Determine unknown values.
What does a pH buffer do?
It controls pH and keeps it at constandt.
What does a colorimetre measure?
Quantify concentration and turbidity.
What does measuring absorbance show?
Allows you to determine the concentration of a coloured solution using suitable wavelength filtres.
What does measuring percentage transmission show?
Allows you to determine turbidity.
What is an example of turbidity
Cells in suspension.
What does a centrifuge do?
Separates substances of
differing density
In a centrifuge where does more dense components settle?
In the pellet
In centrifuge where does less dense components settle?
Supernatant.
What does Paper and Thin layer chromatography do?
It separates different substances such
as amino acids and sugars
What determines the speed of the solute in Paper and Thin layer chromatography
The speed that each solute travels along the
chromatogram depends on its differing
solubility in the solvent used
What do affinity chromatography do?
separate proteins.
How does affinity chromatography work?
- A solid matrix or gel column is created with
specific molecules bound to the matrix or gel. - 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.
What does gel electrophoresis separate?
Proteins and nucleic acids
What happens in gel electrophoresis?
Charged macromolecules move through an
electric field applied to a gel matrix
What do native gel separate?
Proteins by their shape, size and charge.
What happens with native gels?
Native gels do not denature the molecule so
that separation is by shape, size and charge.
What does SDS-PAGE separate?
Proteins by size alone.
What happens in SDS-PAGE?
SDS–PAGE gives all the molecules an
equally negative charge and denatures them,
separating proteins by size alone
What is an Iso-electric point (IEP)?
IEP is the pH at which a soluble protein has
no net charge and will precipitate out of
solution.
What happens to the IEP is a solution is buffered?
If the solution is buffered to a specific pH,
only the protein that have an IEP of that
pH will precipitate
How can proteins be separated using their IEP’s?
Using Electrophoresis
How can electrophoresis separate proteins using their IEP?
Soluble proteins can be separated using an
electric field and a pH gradient. A protein
stops migrating through the gel at its IEP in the pH gradient because it has no net
charge.
What is Immunoassay techniques used for?
To detect and identify specific proteins.
What are used in Immunoassays?
use stocks of antibodies
with the same specificity
What are monoclonal antibodies?
stocks of antibodies
with the same specificity
What is a chemical label used for
linked to an antibody specific to the protein antigen
What is another name for the label and what does it cause?
It is also known as a reporter enzyme
producing a colour change.
What are the two other types of chemical labels used?
Chemiluminescence and fluorescence.
What is Western blotting?
Western blotting is a technique, used after
SDS–PAGE electrophoresis
what happens in western blotting after SDS-PAGE electrophoresis?
The separated proteins from the gel are
transferred (blotted) onto a solid medium. The proteins can be identified using specific
antibodies that have reporter enzymes
attached
What is Bright-field microscopy used for?
used to observe:
- whole organisms,
- parts of organisms,
- thin sections of dissected tissue
- 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 aseptic technique?
Aseptic technique eliminates unwanted
microbial contaminants when culturing microorganisms or cells.
What does aseptic technique involve?
Aseptic technique involves the sterilisation of equipment and culture media by heat or
chemical means and subsequent exclusion of
microbial contaminants.
How can a microbial culture be started?
A microbial culture can be started using an
inoculum of microbial cells on an agar
medium, or in a broth with suitable nutrients
When culturing, what do animal cells grow in?
Animal cells are grown in medium containing
growth factors from serum.
What are growth factors?
Growth factors are proteins that promote cell growth and proliferation. Growth factors are essential for the culture of most animal cells.
What are primary cell lines
primary cell lines can divide a limited number of times,
What does plating out of liquid microbial culture on solid media allow?
allows the number of colony forming units to be counted and the density of cells in the culture estimated
What is required to achieve a suitable colony count?
Serial dilution
What does a haemocytometer do?
estimate cell numbers in a liquid culture
What does vital staining do?
Vital staining is required to identify and count
viable cells
What are the examples of hazards?
- toxic or corrosive chemicals,
- heat or flammable substances,
- pathogenic organisms
- mechanical equipment.
