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    Home > Biochemistry News > Microbiology News > Laboratory disinfection and sterilization technology

    Laboratory disinfection and sterilization technology

    • Last Update: 2021-01-21
    • Source: Internet
    • Author: User
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    sterilization is very important to the research and application of cell and embryo engineering, which directly affects whether the whole experiment can be carried out smoothly.
    (i) Sterilization methods
    Currently commonly used disinfection methods using physical methods (such as dry heat sterilization, wet heat sterilization,
    filtration
    sterilization method, ray sterilization method, etc.) and chemical methods (disinfectants, antibiotics) two categories.
    1. Dry heat sterilization
    is a method of using
    string
    to dry the
    high heat of 120 oC to 150 oC in a box and keeping it for 90 to 120 minutes, killing bacteria and spores for sterilization purposes. It is mainly suitable for the sterilization of glassware, metal instruments and items that cannot be in contact with steam in the pressure steam sterilizer and which are not easily damaged by high temperature. Items sterilized by this method are dry and easy to store. Alcohol lamp flame burning sterilization method is also one of the methods of dry heat sterilization, in the animal cells in-body
    cultilation
    work, often need to use the work surface of the alcohol lamp flame to supplement the metal appliances and glassware mouth edge sterilization.
    2. Wet heat sterilization method
    pressure steam wet heat sterilization method is currently the most commonly used method of sterilization. It uses high-pressure steam and latent thermal action and good penetration in the steam environment to cause the coagulation and denaturation of bacteria
    proteins
    and cause the death of
    microorganisms
    environment. Suitable for the sterilization of cloth work clothes, various utensils, metal instruments, plugs, distilled water, cotton plugs, paper and certain cultures. The steam pressure of the high-pressure steam sterilizer is generally adjusted to 1.0 to 1.1 kg/cm2, and the sterilization effect can be achieved by maintaining 20 to 30 min.
    3. Ray sterilization is
    method of sterilization using ultraviolet light. Ultraviolet rays are low-energy electromagnetic radiation that kills many microorganisms. Ultraviolet rays act by destroying the nucleic acids and proteins of microorganisms. Suitable for laboratory air, ground, operating table sterilization. The sterilization time is 30 min. UV sterilization should be noted, can not be exposed to the side of the experimental operation, because UV not only damage to human skin, but also to culture and some reagents will also have adverse effects.
    4. Filtration sterilization
    is the filtration of liquids or gases through a membrane with micro-pores, so that bacteria larger than the membrane aperture and other microbial particles retain, so as to achieve sterilization methods. Filtration and sterilization method is mostly used in heat easy to break down, denaturation and failure of reagents, enzymes,
    serum
    , culture fluids and so on. At present, the commonly used micro-porous membrane metal filter or plastic filter positive pressure filtration sterilization, or glass bacterial filter, filter ball negative pressure filter sterilization. Membrane aperture should be in the range of 0.22 to 0.45 sm or with a smaller bacterial membrane, after the solution passes through the membrane, bacteria and spores are blocked because they are greater than the membrane aperture, and the use of membrane adsorption, blocking bacteria less than membrane aperture through.
    5. Chemical disinfectant disinfection
    used in items that cannot be sterilized by physical methods, air, work surfaces, operator's skin, certain laboratory utensils, etc. Commonly used chemical disinfectants include formaldehyde, potassium permanganate, 70% to 75% ethanol, peroxyacetic acid, resul water, 0.1% neo-ethylene, ethylene oxide, iodine volts or iodine. Among them, 70% to 75% ethanol, 0.1% to 0.2% mercury chloride, 10% sodium hypochlorate, saturated bleach, etc. were used to sterilize experimental materials; Safety should be taken care when using, especially disinfectants used on skin or experimental materials, and appropriate types, concentrations and treatment times must be selected in order to achieve safety and sterilization purposes.
    6. Antibiotic antibacterial method
    mainly used in culture fluid, is an important means to prevent microbial pollution in the culture process and as a microbial pollution is not serious when the "first aid" measures. Common antibiotics are penicillin, streptomycin and neomycin.
    (ii) The disinfection and sterilization methods of different kinds of articles and cultures
    1. Sterilization of the sterile operating room
    cultured materials when cultured, observed or replaced, any contaminants must be prevented from entering the culture fluid or container in all aspects, so sterilization of the sterile operating room is essential. Since the source of pollution in the sterile operating room is mainly bacteria and fungal spores in the air, the sterile operating room after long-term deactivation should be fumigated and sterilized, fumigation is the sterilization of the sterile chamber with potassium permanganate and formaldehyde (2 ml formaldehyde plus 1g potassium permanganate) / m3. The frequently used sterile operating room should be cleaned on the ground before each use and exposed to 30 min with ultraviolet light for air sterilization. For the ultra-clean work station, use UV lamps to shine 30 min before each operation, then wipe with 70% to 75% alcohol.
    2. Culture liquid sterilization
    culture liquid in the preparation process with a variety of miscellaneous bacteria, should be installed immediately after sterilization, or within 24 hours to complete the sterilization process. At present, filtration and sterilization methods are commonly used to remove bacteria from culture operation fluids and culture fluids. Or the heat-resistant components in the culture first high-pressure steam sterilization, and then in the sterile chamber to add filtered sterilization of the heat-resistant solution, mixed and evenly divided into equipment.
    When using a high-pressure steam sterilizer to sterilize, put the packed culture liquid into a low-pressure steam sterilizer with a certain amount of cold water (inundated heating tube) at the bottom, and pressurize to 0.35 to 0.4 kg/cm2 to drain the cold air inside the sterilizer, so that the steam can reach the various disinfection sites, to ensure thorough disinfection. Then continue to pressurized heating, when the pressure meter reading reaches 1.0 to 1.1 kg/cm2 is 121 oC, to maintain 15 to 20 min.
    sterilization effect depends on temperature rather than pressure, it is necessary to maintain a long disinfection time under certain pressures. At a steam temperature of 121 oC, a variety of bacteria and highly heat-resistant Bacillus spores can be killed quickly, so the disinfection pressure and temperature of many cultures are generally maintained at 1.0 to 1.1 kg/cm2, 121 oC. Sterilization time is closely related to the amount of culture fluid that needs to be disinfected (Table 2-3), the time is not enough to achieve the sterilization effect, some chemical substances in the long-term culture solution are destroyed, affecting the culture solution composition.
    sterilization, close the heat source, only when the pressure in the sterilizer is reduced to zero can open the valve, eliminate the remaining steam, remove the culture solution. Do not open the air valve when the pressure is high, causing the decompression boiling, so that the liquid in the container overflow. If the composition of the culture liquid contains substances that are easily decomposed by heat, sterilization methods are required. First of all, the heat-resistant components in the culture of high-pressure steam sterilization and placed in a sterile place, solid culture liquid in 40 oC or so
    agaride
    is about to condense, add filtered sterilization of the heat-resistant solution, after mixing and cooling standby. Liquid culture is cooled to room temperature before adding a filtered sterilization solution. When filtration of sterilization, a bacterial membrane with an aperture of 0.22 to 0.45 m or less is used. Filtration and sterilization can be carried out using a filtration device or injection filter, the utensils used should be sterilized by high pressure, the temperature should not exceed 121 oC.
    3. Glassware, plastic utensils and instruments sterilization
    glassware can be dried hot or steam-pressure sterilized, but it is best to dry moisture in time after steam sterilization. Plastic utensils that cannot be sterilized by steam at high pressure can be soaked with 75% alcohol and repeatedly sterilized with ultraviolet light while drying on a sterile operating table before use. Laboratory can also use ethylene oxide sterilization bag to disinfect plastic utensils, disinfection of the utensils should be fully dispersed 2 to 4 hours before use. Aseptic operation of a variety of equipment, generally using dry heat or auto-pressure steam sterilization, or 75% alcohol immersion, and then dry on the sterile operating table at the same time with UV repeated sterilization;
    4. Sterilization of cultured materials
    experimental materials taken from animal bodies, carrying microorganisms and impurities, surface disinfection and sterilization must be carried out before vaccination, and materials that have been infected by microorganisms should be eliminated. Some tissue blocks collected from animal bodies shall be soaked with disinfectant and surface disinfected. Common disinfectants have hydrogen oxide (10 to 12%, soaked 5 to 15 min), peroxyacetic acid (0.05%, soaked 30 s to 1 min), alcohol (70 to 75%, soaked 2 min).
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