Distilled water with a resistivity not less than 300,000 Ω·cm or water of the same quality should be used for the preparation of medium used in microbiology, preferably not deionized water produced by ion exchange.
When weighing, special glue should be used to avoid cross-contamination and affect the test results; dry powder culture medium is easy to absorb moisture, if conditions permit, the culture medium should be weighed in a room with low humidity.
When reconstituting various types of culture medium in microbiology, copper or iron pots should not be used to prevent ions from mixing into the medium used in microbiology and affecting the growth of microorganisms; the size of the container should be more than twice the total volume of the cultured medium after reconstitution, to avoid overflow of the culture medium during heating and dissolution; the culture medium containing agar powder can be soaked for a few minutes before heating and dissolving; when heating the culture medium, stirring must be carried out, especially for agar-based culture medium. To avoid burning and boiling over, it is best to use a boiling water bath to heat small amounts of culture medium. Burnt culture medium can destroy nutrients and produce toxic substances. If you find that the culture medium is burned or overflows unevenly before it is completely dissolved, the culture medium cannot be used and should be prepared again. When re-melting agar based medium, a boiling water bath or flowing steam should be used for heating, and the media can only be heated twice at most. The unused culture medium after the second melting should be discarded.
The general culture medium adopts moist heat sterilization, that is, high-pressure steam sterilization, which is usually 121°C for 15 minutes. According to national standards or the instructions provided by the manufacturer, sugars or special culture media should be sterilized. The prepared culture medium must be sterilized immediately to avoid microbial growth and reproduction consuming nutrients, and changing the pH of the culture medium. High temperatures can destroy the nutritional components of the culture medium and also make the gel strength of agar decrease. Therefore, sterilization temperature and time must be strictly controlled, and sterilization cannot be repeated. The pressure gauges of high-pressure steam sterilization pots and other equipment should be checked regularly, and the sterilization effect should be verified by biological indicator or chemical color change paper.
Microorganisms can only grow and reproduce or show their biological characteristics within the appropriate pH range. The pH required by the formula of medium used in microbiology is the pH value after sterilization or disinfection, that is, the pH before the culture medium is used, which should be measured at 25°C using a precision acidity meter. Solid agar culture media should be measured with a special colloidal surface measuring electrode. If it is necessary to adjust the pH of the culture medium during use, attention should be paid not to adjust the pH repeatedly, otherwise it will affect the osmotic pressure of the culture medium.
After sterilization, the culture medium should be cooled rapidly to the required temperature to avoid being stored in the sterilization pot for a long time, resulting in excessive sterilization which affects the nutritional components or selective effects of the culture medium. The amount of prepared culture medium should be just used up within the longest storage period. Culture media containing dyes should be stored away from light. If the poured agar plates are not used up on the day they are prepared, they should be stored in the refrigerator. If the storage time exceeds 2 days, they should be put in a sealed plastic bag; if the prepared broth culture medium is stored for more than two weeks, it should be placed in a screw-capped test tube or other closed test tube or container to prevent evaporation.
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