Experiments with yeast - 2

Direct observation of yeast population growth by counting the number of yeast cells

Background information

If yeast is added to a liquid containing sugar and other nutrients, kept at an appropriate temperature and oxygen supply, it will grow and multiply in number, as long as sufficient nutrients are available.

A counting cell is a special piece of apparatus - an accurately calibrated microscopic counting chamber.

A rafter cell (above) - made of plastic - consists of 1000 (20x50) squares, each 1mm x 1mm, so they are visible to the naked eye. It uses a special glass coverslip which is easy to break and difficult to replace!
A haemocytometer is a similar and more sophisticated (expensive, fragile) piece of apparatus in which the squares are more difficult to find under the microscope, and which necessitates some more complex calculation. There are several types of haemocytometer, developed in order to count (red and white) cells in blood - but they can be used to estimate the number of any microscopic objects in a liquid.

Suggested action

Each bench to work as a team - dilution and microscope/counting chamber techniques need to be followed up by each member of the team, in order for results to be consistent.

Setting up the experiment

Try 1 spatula of yeast per 100 ml of grape juice in each of 2 500ml beakers, placed in an incubator at 25°C
- one with a thin layer of liquid paraffin
- one without
There is in fact an easier better way to do this!

Perform the dilution and counting exercises immediately after setting up the experiment, and then at regular intervals afterwards - daily or twice daily - for at least a week.

Decisions needed:

What factors are to be kept constant (and how) ?

How to organise team work to gather results.

Individually, think about the problem and try to come up with a hypothesis about the pattern of population growth likely to be shown by each culture of yeast. Justify this with appropriate background statements.

Method of counting the yeast population

1) Dilution (may not be required at the start of the experiment, or may need modification or repetition as the experiment proceeds)

Ensure the contents of the beaker are thoroughly mixed - use stirring rod or pipette. Using plastic bulb pipette, remove some yeast suspension from the beaker (below the liquid paraffin layer). Add 1ml of this yeast suspension to a measuring cylinder with a lid, make up to 100 ml and mix by inverting. This will dilute the yeast suspension by 100 times. It may be necessary to repeat the process using a new pipette and measuring cylinder if the result is too concentrated, and the dilution factor must be remembered.

2) Using the counting chamber

Using a different pipette (if diluting as above), transfer 2-3 drops of the diluted suspension to the centre of a "Rafter cell" slide, placed on paper towel, and carefully lower on the special coverslip. Mop up any excess.
Place the counting chamber on the microscope, and examine carefully on low power. Focus very carefully, reducing light if necessary - look for horizontal and vertical lines.

Focus carefully, then move to higher power. Yeast cells are round or oval dots, slightly above the lines marked on the chamber. Do not confuse these with air bubbles, dust or other debris above or below the coverslip or counting chamber.

Perform several counts of the number of yeast cells within the squares, and record your raw data in an efficient way.

Count any buds you see as separate cells.
To avoid counting some cells twice, do not include yeast cells touching "north" or "east" sides of squares.

The yeast population density is given by the following:

Number of cells per ml =

average count per square x 1,000,000 x 100
(or whatever dilution factor is used)

After use, return the microscope to low power.

CAREFULLY wash up the counting chamber AND COVERSLIP, wipe them and place them away in their box.

Put pipettes and measuring cylinders into the washing-up bowl

Results format

These results will be best presented as a graph, eventually.

Day	Time	Elapsed time from start	Counts from beaker 1 - surface covered	Counts from beaker 2 - surface uncovered	Reading taken by (name) :