4.4: Turbidity

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As seen in Lab 2, when you mix the bacteria growing in a liquid medium, the culture appears turbid. This is because a bacterial culture acts as a colloidal suspension that blocks and reflects light passing through the culture. Within limits, the light absorbed by the bacterial suspension will be directly proportional to the concentration of cells in the culture. By measuring the amount of light absorbed by a bacterial suspension, one can estimate and compare the number of bacteria present.

The instrument used to measure turbidity is a spectrophotometer. (See Fig. \(\PageIndex{1}\).) It consists of a light source, a filter which allows only a single wavelength of light to pass through, the sample tube containing the bacterial suspension, and a photocell that compares the amount of light coming through the tube with the total light entering the tube.

Photograph of a spectrophotometer. — Figure \(\PageIndex{1}\): **A Spectrophotometer.** (Copyright; Gary E. Kaiser, Ph.D. The Community College of Baltimore County, Catonsville Campus CC-BY-3.0)

The ability of the culture to block the light can be expressed as either percent of light transmitted through the tube or the amount of light absorbed in the tube. (See Fig. \(\PageIndex{2}\).) The percent of light transmitted is inversely proportional to the bacterial concentration. (The greater the number of bacteria, the lower the percent light transmitted.) The absorbance, or optical density, is directly proportional to the cell concentration. (The greater the number of bacteria, the higher the absorbance.)

Illustration of the concept of a spectophotometer. — Figure \(\PageIndex{2}\): **Illustration of the Concept Behind a Spectrophotometer** (Copyright; Gary E. Kaiser, Ph.D. The Community College of Baltimore County, Catonsville Campus CC-BY-3.0)

Turbidimetric measurement is often correlated with some other method of cell count, such as the direct microscopic method or the plate count. In this way, turbidity can be used as an indirect measurement of the cell count. For example:

1. Several dilutions can be made of a bacterial stock.
2. A Petroff-Hausser counter can then be used to perform a direct microscopic count on each dilution.
3. Then a spectrophotometer can be used to measure the absorbance of each dilution tube.
4. A standard curve comparing absorbance to the number of bacteria can be made by plotting absorbance versus the number of bacteria per cc. (See Fig. \(\PageIndex{3A}\).)
5. Once the standard curve is completed, any dilution tube of that organism can be placed in a spectrophotometer and its absorbance read. Once the absorbance is determined, the standard curve can be used to determine the corresponding number of bacteria per cc. (See Fig. \(\PageIndex{3B}\).)

Fig. \(\PageIndex{3A}\): A Standard Curve Plotting the Number of Bacteria per cc versus Absorbance, Step 1	Fig. \(\PageIndex{3B}\): Using a Standard Curve to Determine the Number of Bacteria per cc in a Sample by Measuring the Sample's Absorbance, Step 2

(Copyright; Gary E. Kaiser, Ph.D. The Community College of Baltimore County, Catonsville Campus CC-BY-3.0)

Contributors and Attributions

Dr. Gary Kaiser (COMMUNITY COLLEGE OF BALTIMORE COUNTY, CATONSVILLE CAMPUS)