PAG - Using aseptic technique to investigate the use of antimicrobials Flashcards
antibiotics
used to treat bacterial infections
how can we kill bacteria using antibiotics
- interfering with building of bacterial cell wall by inhibiting synthesis of peptidoglycan
- preventing bacteria from multiplying by interfering with DNA replication preventing cell division
what can discs of paper soaked in antibiotic be tested for
- how effective they are in treating bacterial infections
the greater the zone of inhibition …
the more effective the antibiotics are at killing that bacterium
Health and Safety - list
- bunsen burners
- ethanol
- e -coli
- agar plates
- after experiment
bunsen burners
- use yellow flame when not in use
- tie up long hair
- wear eye protection
ethanol
- may be used
- equipment placed in ethanol, e.g inoculating loop/spreaders need to be place in flame to burn off ethanol
- this keeps the equipment sterile
- dispose equipment
e - coli
- despite strain being safe, there is a small chance of infection
- place equipment in disinfectant after use
- wash hands before and after use
agar plates -
- should not be completely sealed
- this is to prevent hazardous anaerobic bacteria from growing
- only close with 3 pieces of sticky tape
after experiment
- once inhibition zones have been measured
- clean hands, clean desk
- place equipment in disinfectant
First step - preparation
- wash hands
- soak a piece of cotton wool in disinfectant jug and thoroughly clean work area
- label agar plate with species of bacteria around the edges
- turn agar plate over to do this
Second step - bunsen burner
- light Bunsen burner
- turn to blue flame
- pick up a sterile pipette in dominant hand and place between thumb and first finger
- lift bottle of e-coli to dominant hand and open bottle
- pass neck of the bottle through the flame and use a sterile pipette to draw 0.5cm3 of e-coli
- flame neck of bottle again
- turn Bunsen burner back to yellow flame
After we have drawn 0.5cm3 of e coli what do we do
- lift one side of petri dish towards BB
- use pipette to squeeze e-coli in
- place lid back on immediately
- put pipette in disinfectant
after we have put e-coli onto agar plate what do we do with a spreader
- take a sterile spreader
- evenly spread e-coli culture across the surface of the agar plate
- place the lid back on and put the spreader in disinfectant
after spreading the e coli evenly using a spreader - what do we do (start with bunsen)
- turn blue flame
- sterilise the edges of forceps
- lift agar plate lid and place mast ring onto surface of agar in the centre
- use forceps to ensure every antibiotic disc is in contact with gel
after placing down the mast ring what do we do
- place lid on agar plate
- put forceps in disinfectant
- secure lid using 3 pieces of tape
- place plate with jelly base downwards onto tray
- incubate for 24 hours
after incubation for 24 hours, what do we do
- use cotton wool and disinfectant to clean desk
- wash hands
- examine plates but don’t remove lid
- keep plate upside down and use a ruler to measure inhibition zone
- draw rings of inhibition on diagram and put measurements into a table
- place plates back onto tray so they can be autoclaved
why did we pass the neck of e coli culture through a blue bunsen flame
- to sterilise it
- kill microorganisms surrounding the air in the bottle
- air in the bottle expands = outflow of air out so bacteria couldn’t enter bottle
why wasn’t the lid of the agar plate completely sealed with tape
- it needs some air/oxygen to respire aerobically
why are agar plates incubated upside down
- so water vapour doesn’t end up on the lid
- this could affect bacteria growth
- water vapour would condense on agar = pathogenic
Control experiment for investigation
- have another disc with no antibiotic to see how bacteria grows
reproducible
different scientists can follow the same procedure and find the same trends
repeatable
same scientist does the same method and gets the same trends
how do we make a method reproducible
- make it valid
- variables should be controlled
