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Alevel Homework Help

I have biology homework that I seriously need help with. Any help at all is super appreciated :smile:

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"Estimate the number of bacteria in milk (ATc & ATi)

Research a method that could be used to count the num. of bacteria in a sample of milk. The problem is that the broth will contain too many bacteria to count using a microscope.

Your method should use standard laboratory glassware and equipment.

How can we use agar plates to enable us to count the number of bacteria in the original milk sample? You should refer to aseptic technique throughout the plan produced.

Hint: Serial Dilution"

Reply 1

JessNaomi19600

I'm doing a-level biology and I recently did this as a practical experiment. You have to carry out a serial dilution. Here is the process we used:
1) Firstly, sterile workplace must be created by wiping the worksurface with an antibacterial spray.
2) Set up five sterile, 25ml beakers and add 9.9cm of sterile, distilled water to each, using a 10ml sterile syringe.
3) Make sure you label each dilution to keep track of the concentration of each beaker: 10^-2, 10^-4, 10^-6, 10^-8 and 10^-10.
4) Then, add a 0.1cm³ sample of the milk to the first sample (labelled 10^-2), using a sterile 1cm³ syringe.
5) Next, mix the 10^-2 solution and, using another sterile 1cm³ syringe, add a 0.1cm³ sample from this solution, to the next beaker (labelled 10^-4).
6) Repeat this until you obtain a solution for each concentration (up to 10^-10).
7) Set up a sterile environment by setting up a Bunsen burner next to your workspace. This creates a sterile field.
8) Flame a loop by holding in the Bunsen flame and then dip it into the sample. Then, spread the loop gently over the agar in an agar plate, only taking the lid of the agar plate of halfway to ensure no other bacteria enter. Place the lid back down quickly, tape it down at the edge, label it with its concentration and place it in an incubator for 72 hours
9) Repeat this for each concentration, flaming the metal loop after each and place in the incubator.

After the three days, remove the agar plates from the incubator and analysis them. The colonies of bacteria in the sample should be visible in each agar plate. However, the more dilute solutions would have produces less bacteria and so you can distinguish single colonies.
You should pick an agar plate which has a viable count (a number of colonies which can be counted). You should not pick an agar plate that has too many bacteria as the colonies will not be distinguishable. You should also not choose an agar plate with too few bacterial colonies because their won't be enough bacteria to obtain an accurate, reliable result.
Once you have counted the number of colonies, you should record this number and times it by the dilution factor to obtain an estimate of the number of bacteria in the original sample of milk.