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Microbiology Concept Inventory
Please note, you must be an educator in higher ed or maybe high school to qualify to recieve the MCI
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|Counting chambers
The most direct method of counting microorganism is by the use of a microscope and a slide with special chambers of known volume. These slides allow the counting of a small number of cells in a small volume and extrapolating the result to determine the population. An example of such a device is shown in Figure 4-10. A culture is placed on the slide marked with precise grids. The number of cells present in each grid is counted and an average determined. Conversion using a formula gives the number of cells per milliliter in the culture. This method is rapid, a result can be known in just a few minutes, and is easy to perform. However, it is impossible to distinguish living cells from dead ones. If this distinction is important, direct microscopic counts are not the solution. Finally, cultures containing less than 1 million cells per ml are actually too dilute for direct counts since there will be too few cells in the very small volume that is actually examined under the microscope for an accurate count.
Figure 4.10. The Petroff-Hauser counting chamber. The center of the slide contains a precisely machined grid, with each square (1/20 mm x 1/20 mm) having a known area. The coverslip also rests above the slide a known distance (typcially 1/50 of a mm). Since each of these dimenstions is known, it is possible to calculate the number of cells in each square and pluggin it into a formula. On the left of the figure is a photograph of a Petroff-Hauser slide. On the right is a grid at 100 x magnification showing the size of the squares.
Electronic particle counters
Electronic particle counters are useful if the number of bacteria in a sample needs to be counted on a routine basis. The method is based on the property that nonconductive particles, such as bacteria, will cause a disruption in an electric field as they pass through it. A Coulter counter is a type of electronic particle counter in which there is a small opening between electrodes through which suspended particles pass, see Figure 4-11. In this sensing zone, each particle displaces its own volume of electrolyte, causing a current pulse. The pulse is noted and recorded as one particle count. By precisely controlling the rate at which solution passes through the opening, it is possible to get exact, reproducible counts at a rate of up to several thousand bacteria per second. Coulter counters are highly dependent upon particle size and those dependent upon changes in current are near their detectable limits with microorganisms. Particle counters that use light diffraction as a means of sizing and counting particles are also manufactured and can detect particle less than 1 µm in diameter.
Figure 4.11. The coulter counter. A picture of a Coulter Counter. The diagram at the right demonstrates the opening that the microbes must pass through during counting. When microbes pass through the aperture the electrical potential across the electrodes is distrubes, which the dataprocessing system records as a count. In the lower right is an actual picture of a Multisizerâ„¢ 3 by Beckmann instruments that uses the Coulter counter priciple.
The advantage of this method is the simplicity of its operation and it reproducibility. As in microscopic counts, the machine cannot distinguish between living or dead cells or even between dust and bacteria. Any reasonably sized particle in the solution will be counted. There is also the expense of buying the counter, which can cost many thousands of dollars.