Serial Dilution

• Place 9cm3 of distilled water into test tubes with bungs.
• Place 1cm3 of the bacterial sample in the first tube and mix.
• Transfer 1cm3 of solution from the first tube into the next tube and mix, repeat for the remaining tubes.
• Transfer 1cm3 of a sample from each tube onto a sterile nutrient agar plate.
• Use a sterile spreader to spread the sample around each plate.
• Repeat this so there is 3 plates per dilution (allows calculation of an average number of colonies)
• Seal with tape but not air tight (around the plate) this prevents growth of anaerobic bacteria.
• Incubate at 25'C for 24hours, don't incubate at 37'C as this is optimum for human pathogenic bacteria.
• Count number of colonies on each plate.
• Multiply the number of colonies by the diultion factor to give the number of bacteria in the original sample. 

Serial diultion = a viable count that assumes a single bacterial cell will reproduce to form a  colony that can be seen and counted. 

Inaccuracies with experiment:

• Clumping - can lead to an underestimation
• Every dilution decrease accuracy 
• Count the lowest dilution with no clumping