Summary of methods for determining the number of bacteria

1. The color change unit (CCU)
This method is usually used for microorganisms that are small and cannot be counted by general turbidimetry, such as mycoplasma, because the liquid culture of mycoplasma is completely transparent and appears clear and transparent red, so it cannot be counted by turbidimetry. Since the solid culture of mycoplasma is difficult, it is not easy to count by the cfu method, so a special counting method, that is, the CCU method is required. It uses the metabolic activity of microorganisms in the medium as an indicator to count the relative content of microorganisms. The following is an example of Ureaplasma urealyticum, and its operation is briefly described:
(1) Take 12 sterile test tubes, each tube containing 1.8 ml of Ureaplasma urealyticum.
(2) Add 0.2ml of ureaplasma urealyticum solution to be tested in the first tube, mix well, draw 0.2ml from the second tube, and so on, 10 times gradient dilution, until the last tube (3) At 37 degrees, the last tube with the color change of the medium is used as the CCU of the test solution, that is, the maximum metabolic activity of the mycoplasma. For example, the sixth tube has a color change, and its relative concentration is 10 to the 6th power CCU. /ml.
In general, turbidimetry and colony counting can satisfy the counting of most bacteria, but for the special microorganisms such as mycoplasma, the CCU method is more suitable.
There are many methods for determining the growth of microorganisms, and various methods have their advantages and disadvantages, and are not applicable in any case. Commonly used in microbiology work are the plate colony counting method, the counter method and the turbidimetry method. As for which method is more suitable for you, it depends on your specific conditions.
2. Counter measurement method:
That is, counting with a blood cell counter. A volume of sample cell suspension was placed in the counting chamber of the hemocytometer and counted by microscopic observation. Since the volume of the counting chamber is constant (0.1 mm3), the number of bacteria in the sample can be calculated based on the number of bacteria in the counter scale. This method is simple and easy to use and results are immediately available.
This method is not only suitable for bacterial counting, but also for yeast and mold spore counting.
3, electronic counter counting method:
The working principle of the electronic counter is to measure the change of the resistance of the liquid in the small hole. The small hole can only pass through one cell. When a cell passes through the small hole, the resistance increases obviously, forming a pulse, which is automatically recorded on the electronic recording device.
The method is more accurate, but it only recognizes the particle size and does not distinguish whether it is a bacterium. Therefore, it is required that the bacterial suspension does not contain any debris.
4. The live cell counting method commonly used is a plate colony counting method, which is designed according to the principle that each living bacteria can grow a colony. Take a certain volume of the bacterial suspension, make a series of dilutions, and then plate the quantitative dilution, and calculate the number of viable cells in the culture according to the number of colonies cultured. This method has high sensitivity and is a method for detecting the number of live bacteria in pollution. It is also a method adopted by many countries in the world. Use this method should pay attention to: 1 generally select the number of colonies between 30 ~ 300 to count, too much or too little are not accurate; 2 in order to prevent colony spread, affect the count, can be added to the medium O. 001% 2,3,5 triphenyltetrazolium chloride (TTC); 3 This method is limited to microorganisms forming colonies.
It is widely used in bacteria testing of various materials such as water, milk, food, medicine, etc. It is the most commonly used live bacteria counting method.
5. Turbidimetric turbidimetry is the indirect determination of the number of bacteria based on the amount of light transmitted from the bacterial suspension. The concentration of the bacterial suspension is inversely proportional to the transmittance within a certain range and is proportional to the optical density. Therefore, the bacterial liquid can be measured by a photoelectric colorimeter, and the optical density (OD value) is used to indicate the concentration of the sample liquid.
This method is simple and quick, but it can only detect suspensions containing a large number of bacteria, and the relative number of bacteria is obtained. For samples with too dark color, this method cannot be used.
6. Determination of cell weight method This method is divided into wet weight method and dry weight method. The wet weight method is used to weigh the wet cells in a unit volume culture after centrifugation; the dry weight method is centrifuged, and the mixture is washed with water and dried by a dryer to lose moisture and then weighed. .
This method is suitable for samples with high bacterial concentration and is a commonly used method for determining the growth of filamentous fungi.
7. Determination of total nitrogen or total carbon content of cells. Nitrogen and carbon are the main components of cells, and the content is relatively stable. The determination of nitrogen and carbon content can infer the quality of cells. This method is suitable for samples with high cell concentrations.

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