What you need to know
- 6.2Cell counts, dilution platingSpreading a known volume of a diluted culture on agar and counting colonies, to estimate the number of viable cells., mass and optical (turbidityCloudiness of a liquid culture. It increases with the number of cells and can be measured with a colorimeter.) methods.
k from those numbers is 6A 3. Liquid-culture method and ethics are CP13. Colourimeter brand names are not required.
By the end, you should be able to…
- List the four specified methods.
- Distinguish a total countA count of all cells, living and dead, for example with a haemocytometer. from a viable countA count of living cells only, for example from colonies grown on agar..
- Explain why turbidity and mass can overestimate living cells.
Build your understanding
Total versus viable
A cell count with a counting chamber records numbers in a known volume, living and dead: a total countA count of all cells, living and dead, for example with a haemocytometer.. dilution platingSpreading a known volume of a diluted culture on agar and counting colonies, to estimate the number of viable cells. serially dilutes the culture, spreads a known volume on agar, counts colonies after incubation, and multiplies by the dilution factor to give colony-forming unitA viable cell or clump of cells that grows into one colony on agar. Counts are given as cfu per cm³. per cm³: a viable countA count of living cells only, for example from colonies grown on agar..
serial dilutionA series of dilutions, each by the same factor (often 10), used to get a countable number of cells. exists so that colonies are separate enough to count, and so that you can scale back to the original density. One colony is treated as one CFU, which assumes each colony grew from one cell — a stated assumption, not a photograph of that cell.
Mass and turbidity
Mass, especially dry mass, measures harvested biomassThe mass of living material, often measured as dry mass per unit area. and avoids variable water in wet mass. It includes dead material and destroys that sample. turbidityCloudiness of a liquid culture. It increases with the number of cells and can be measured with a colorimeter. uses a colorimeter: more cells scatter more light, so absorbance of the mixed broth rises. It is fast and can be repeated, but dead cells still scatter light.
Mix the culture before each optical or count sample so settled cells do not bias the reading. A blank of sterile medium sets the zero. Absorbance is not itself a cell number until you calibrate, and even then it is not a viable count.
Choose for the question
If the question wants living cells, dilution plating is the method that matches. If it wants a quick time course in one flask, turbidity is practical and must be evaluated. If it wants biomass, mass is honest about water. If it wants a microscope number today, a chamber count will do and will include corpses.
Fictional plating example: 62 colonies from 0.10 cm³ of a 10⁻⁴ dilution. CFU cm⁻³ = 62 × 10 × 10⁴ = 6.2 × 10⁶. Those numbers are for practising the scale factor, not a result to copy.
NOT: only dilution plating estimates living colony-forming units. The other three include dead cells or non-living biomass.
- Worked plate: 62 colonies from 0.10 cm³ of a 10⁻⁴ dilution
- CFU cm⁻³ = 62 × 10 × 10⁴ = 6.2 × 10⁶
- One colony is assumed to come from one cell
- Mix the culture and zero the colorimeter with sterile medium
- Absorbance is not a cell number until it is calibrated
Two traps this diagram blocks
- Do not call a colorimeter reading a count of living cells.
- Do not forget to scale both the volume and the dilution.
From the diagram alone
Question
62 colonies grow from 0.10 cm³ of a 10⁻⁴ dilution. Calculate the number of CFU per cm³ and state what this method counts that a turbidity reading does not. [4]
Show a model answer
Per cm³ the colony count is multiplied by 10, and the dilution factor is 10⁴, so CFU cm⁻³ = 62 × 10 × 10⁴ = 6.2 × 10⁶. Plating estimates viable cells. Turbidity also records dead cells, because they still scatter light.
| Method | Living only? | A limitation |
|---|---|---|
| Cell count | No | Dead cells included; small volumes |
| Dilution plating | Yes (CFU) | Slow; assumes one cell per colony |
| Mass | No | Includes dead biomass; destructive |
| Turbidity | No | Dead cells still scatter light |
Make your reasoning visible
Say viable or total
“Count” alone is ambiguous.
Show the dilution arithmetic
Colonies × (1 / volume plated) × dilution factor.
Key explanations
- The calculation assumes one colony grew from one cell. Clumps break that assumption.
Avoid this misconception
The claim: All four methods count living bacteria.
The correction: Only dilution platingSpreading a known volume of a diluted culture on agar and counting colonies, to estimate the number of viable cells. estimates viable CFUs. Cell counts, mass and turbidityCloudiness of a liquid culture. It increases with the number of cells and can be measured with a colorimeter. include dead cells or non-living biomassThe mass of living material, often measured as dry mass per unit area..
How many living bacteria are in a drop of broth?
A broth culture can hold millions of bacteria in every cm³: far too many to count directly. Diluting it in steps and growing a small sample on agar turns that into a number you can count.
What the model shows
A made-up broth culture, a series of tenfold dilutions, and agar plates spread with 0.10 cm³ from the tubes you choose. After incubation each living cell has grown into a colony.
What to look for
Which plates have too many colonies to count, which have too few, and which one falls in the 30 to 300 range that you can count reliably.
What it tells us
Counting the colonies on one good plate and multiplying back by the volume and the dilution factor estimates the living cells in the original broth, in colony-forming units per cm³.
Be careful what you claim
It assumes each colony grew from one cell, and the count varies by chance from plate to plate. Dead cells never form colonies, so this is a viable count, not a total count.
Try it in the model: make all six dilutions, spread plates from 10⁻³, 10⁻⁴ and 10⁻⁵, then incubate them.
Serial dilution and plate counts
A fictional broth holds millions of bacteria per cm³. Dilute it tenfold six times, spread 0.10 cm³ from the tubes you choose, incubate, then count a plate with 30–300 colonies and scale back to CFU per cm³.
What you see
- A broth culture and six dilution tubes, 10⁻¹ to 10⁻⁶, each shaded by its cell density.
- An agar plate under each tube once it is spread, and the colonies after incubation.
- Plates marked as too many to count, countable (30–300) or too few.
How to use it
- Make the series one transfer at a time, choose which tubes to plate, then incubate.
- Pick a countable plate and scale its count back to CFU per cm³ of the broth.
- Start a new broth to practise with a different density.
What it shows
- Why serial dilution is needed: separate colonies, and a known factor to scale back by (6.2).
- How colonies × (1 / volume) × dilution factor gives CFU cm⁻³.
- That dilution plating is a viable count, unlike a counting chamber, mass or turbidity.
Limits of the model
- Fictional broth and numbers. Each colony is assumed to grow from one cell; clumps make plating underestimate cells.
- Colony numbers include chance (Poisson) variation. Real practice uses replicate plates and averages them.
- Pipetting error, and cells that cannot grow on this medium, are left out.
Guided tour · step 1 of 7
- In the simulator, mix the broth, then transfer 1 cm³ into 9 cm³ of sterile water, six times, to make 10⁻¹ to 10⁻⁶.
- Or press Make the rest in Dilute and plate, just below these steps.
Each tenfold step divides the density by 10. The original broth is far too dense to give separate colonies.
Dilute and plate
Count and scale back
Spread at least one plate and incubate to see colonies.
Questions
Lesson example (fictional): 62 colonies grew from 0.10 cm³ of the 10⁻⁴ dilution. What is the density of the original culture in CFU cm⁻³?
Why 30–300?
Above about 300 colonies, neighbours merge and compete, so counts are too low and hard to make. Below 30, a few chance cells make a big percentage difference. The undiluted broth would give a lawn, which is why the series is needed. Turbidity and dry mass (also 6.2) are faster but include dead cells.
Check your understanding
Try each question before opening the suggested answer.
Check your understanding (continued)
Practice question
Retrieval and explanation · suggested answer
List the four specified methods of measuring microbial growth.
Show suggested answer
Cell counts, dilution plating, mass, and optical methods (turbidity).
Practice question
Retrieval and explanation · suggested answer
Compare a total cell count with a viable count.
Show suggested answer
A chamber count includes living and dead cells. Dilution plating counts cells that form colonies, so it is a viable count.
Practice question
Retrieval and explanation · suggested answer
Why can turbidity overestimate living numbers?
Show suggested answer
Dead cells still scatter light, so absorbance stays high after cells die.
Practice question
Retrieval and explanation · suggested answer
Why use dry mass rather than wet mass?
Show suggested answer
Dry mass removes variable water, so the figure better reflects cell material.
Take the idea further
Apply the same specification knowledge to an unfamiliar problem. Any extra assumptions are supplied in the question.
Stretch question
Fictional data: absorbance stays high while CFU cm⁻³ falls. Interpret, and say which 6.3 phase this pattern fits.
Show a hint
What each method includes.
Explore the reasoning
Cells are dying but still scattering light, so turbidity lags the viable count. That mismatch is expected as a culture enters death phase. k must not be calculated from the absorbance plateau.
Recall this lesson
Lesson recall questions
Question 1 of 5: Compare a total cell count with a viable count from dilution plating.
Measuring microbial growth
Practice question
Retrieval and explanation · suggested answer
Compare a total cell count with a viable count from dilution plating.
Lesson: 6A 5: Measuring the Growth of Bacterial Cultures
Try this from memory first.
Put your biology to work
Explore where these ideas appear in the world. Enrichment, not extra required recall.
Why a brewery still plates as well as reading a meter
6A · Measuring the Growth of Bacterial CulturesThe Microbiology Society treats culture and counting as basic microbial work. A turbidity meter is quick on a tank; a plate count still asks how many cells can…Specification 6.2Read & use the scienceClose article
Why this matters
The Microbiology Society treats culture and counting as basic microbial work. A turbidity meter is quick on a tank; a plate count still asks how many cells can grow. School colorimeter curves are the same distinction at flask scale. Factory numbers are not your CP13 results. Optical density records how much light a suspension scatters; it does not identify live cells. A colony-forming count measures cells or clumps able to grow under the chosen plate conditions, so the two values need not agree.
Back to the specification
Pearson Edexcel IAL Biology · Issue 2, February 2021 · Unit 4, pp. 29–32. The application extends beyond required recall.
- 6.2 understand the different methods of measuring the growth of microorganisms, as illustrated by cell counts, dilution plating, mass and optical methods (turbidity)
Use the science
If a meter reading is convenient, why might an examiner still ask for dilution plating?
Explore a possible response
The question may need a viable count. Convenience does not turn turbidity into CFU.
Explore the source
- Microbiology Society: what is microbiologyMicrobiology Society
Cloudy is not the same as alive
6A · Measuring the Growth of Bacterial CulturesOptical density estimates how much light a microbial suspension scatters. A published study found that stress and reduced viability changed the relationship between optical density and cell count…Specification 6.2Read & use the scienceClose article
Why this matters
Optical density estimates how much light a microbial suspension scatters. A published study found that stress and reduced viability changed the relationship between optical density and cell count in Listeria. A food microbiologist therefore calibrates an instrument for the organism and conditions being tested, and may check a culture by dilution plating. Dead cells and particles can still scatter light; a colony count instead records units able to grow on the chosen medium. Neither method is a perfect census, because clumps and growth conditions affect the result. The lesson’s methods answer different questions, even when both produce a number.
Back to the specification
Pearson Edexcel IAL Biology · Issue 2, February 2021 · Unit 4, pp. 29–32. The application extends beyond required recall.
- 6.2 understand the different methods of measuring the growth of microorganisms, as illustrated by cell counts, dilution plating, mass and optical methods (turbidity)
Use the science
If a disinfectant kills many cells without immediately clearing the suspension, how might turbidity and viable plate count differ?
Explore a possible response
Turbidity may remain high because dead cells still scatter light, while the viable colony count should fall.
Explore the source
- Optical density and cell count under stressInternational Journal of Food Microbiology · Abstract