As bacteria multiply in a liquid broth, the broth becomes cloudy, or turbid. The more cells there are, the more light they absorb and scatter. Turbidity can therefore be used as an indirect measure of cell number.Turbidity is measured with a colorimeter (or spectrophotometer):
1. Put sterile broth in a cuvette and use it as a blank to set the colorimeter to zero absorbance.
2. Place a well-mixed sample of the culture in a clean cuvette and measure its absorbance (or transmission).
3. Repeat at intervals to follow growth. The higher the absorbance, the more cells are present.An absorbance reading does not directly give a number of cells. To convert it, a calibration curve is made first: absorbance is measured for a set of cultures whose cell numbers are known, found by another method such as a haemocytometer count or dilution plating. Absorbance is plotted against cell number, and the cell number of an unknown sample is read from the curve.
Worked example: on a calibration curve, an absorbance of 0.40 corresponds to 2 × 108 cells per cm3. A sample with an absorbance of 0.40 therefore contains about 2 × 108 cells per cm3.Turbidity is quick and does not use up the sample, so readings can be taken many times. However, it is a total count: dead cells make the broth cloudy too. It is also only accurate over the range where absorbance increases steadily with cell number, so very dense cultures must be diluted first.
1. Put sterile broth in a cuvette and use it as a blank to set the colorimeter to zero absorbance.
2. Place a well-mixed sample of the culture in a clean cuvette and measure its absorbance (or transmission).
3. Repeat at intervals to follow growth. The higher the absorbance, the more cells are present.An absorbance reading does not directly give a number of cells. To convert it, a calibration curve is made first: absorbance is measured for a set of cultures whose cell numbers are known, found by another method such as a haemocytometer count or dilution plating. Absorbance is plotted against cell number, and the cell number of an unknown sample is read from the curve.
Worked example: on a calibration curve, an absorbance of 0.40 corresponds to 2 × 108 cells per cm3. A sample with an absorbance of 0.40 therefore contains about 2 × 108 cells per cm3.Turbidity is quick and does not use up the sample, so readings can be taken many times. However, it is a total count: dead cells make the broth cloudy too. It is also only accurate over the range where absorbance increases steadily with cell number, so very dense cultures must be diluted first.