What are the commonly used sterilization methods for microbiology experiments are generally divided into two types: moist heat sterilization and dry heat sterilization.
A, moist heat sterilization method refers to the use of saturated water vapor, boiling water or circulating steam sterilization method, due to the steam latent heat, strong penetration, easy to denaturation or coagulation of proteins, so the sterilization efficiency of this method is higher than that of the dry heat sterilization method, is the most commonly used sterilization method. Moisture-heat sterilization method can be divided into: boiling sterilization, pasteurization, autoclaving, circulation steam sterilization, and intermittent steam sterilization.
(1) boiling sterilization method: boil the water to 100 degrees Celsius, keep 5-10 minutes to kill the bacterial propagules, keep 1-3 hours to kill the germ cells. Add one percent to two percent of sodium bicarbonate in the water when the boiling point of up to 105 degrees Celsius, can enhance the sterilization effect, but also can be decontaminated to prevent rust. This method is applicable to eating utensils, knives and arrows, slides and syringes.
(2) pasteurization: a low-temperature disinfection method, named for Pasteur first. There are two specific methods, one is a low-temperature maintenance method: 62 degrees Celsius to maintain 30 minutes; the second is a high-temperature instantaneous method: 75 degrees Celsius role 15-30 seconds. This method is applicable to the sterilization of food.
(3) circulation steam sterilization method: the use of circulation steam at atmospheric pressure for sterilization.
(4) intermittent steam sterilization method
(5) high-pressure steam sterilization method: 103.4 kPa steam pressure temperature up to 121.3 degrees Celsius, maintained for 15-20 minutes.
Two, dry heat sterilization method refers to the technology of sterilization in a dry environment (such as flame or dry hot air). There are generally flame sterilization method and dry hot air sterilization method.
(1) flame sterilization: refers to the sterilization method of direct flame burning. The method of sterilization rapid, reliable, simple, suitable for flame-resistant materials (such as metal, glass and porcelain, etc.) items and utensils sterilization, not suitable for sterilization of drugs.
(2) dry hot air sterilization method: refers to the method of sterilization with high-temperature dry hot air. The method is suitable for high-temperature-resistant glass and metal products as well as oils and fats (such as oily ointment mechanism, injection oils, etc.) and high-temperature-resistant powder chemicals sterilization, is not suitable for rubber, plastics, and most of the sterilization of medicines.
What are the methods of sterilization commonly used in clinical practice? A physical method
1, temperature
The use of temperature for sterilization, disinfection or antiseptic, is the most commonly used and convenient and effective method. High temperature can make the microbial cell protein and enzyme denaturation and inactivation, so as to play a sterilization effect, low temperature is usually bacteriostatic effect.
1) dry heat sterilization method:
a. Scorching sterilization method: the use of flame directly to the microbial death. This method is thorough and reliable, rapid sterilization, but easy to burn items, so the use of limited scope, only suitable for inoculation needles, rings, test tube mouth and can not be used to contaminated items or the bodies of experimental animals and other sterilization.
b. dry hot air sterilization method: this is a commonly used method in the laboratory, that is, the items to be sterilized evenly into the oven, warmed to 160 ° C, constant temperature for 1 hour can be. This method is applicable to the sterilization of glass dishes, metal utensils and so on.
2) Moisture-heat sterilization method:
In the same temperature, the effect of moisture-heat sterilization is better than dry heat sterilization, this is because on the one hand, the protein in the cell contains high water content, easy to denature. On the other hand, high-temperature water vapor has a high degree of penetration of proteins, thus accelerating protein denaturation and rapid death.
a. Pasteurization: some food will be high temperature to destroy the nutrients or affect the quality, such as milk, soy sauce, beer, etc., so can only be used at lower temperatures to kill the pathogenic microorganisms therein, so as to maintain the nutrition and flavor of the food, but also disinfected to ensure food hygiene. The method is generally at 62 ° C, 30 minutes can achieve the purpose of sterilization. This method is the first French microbiologist Pasteur, so it is called pasteurization.
b. Boiling disinfection method: directly to be disinfected items into the water, boiling for 15 minutes, you can kill all the nutrients and part of the bacterial spores. If you add 1% sodium carbonate or 2% carbolic acid in the water, the effect is better. This method is applicable to the sterilization of syringes, towels and anatomical utensils.
c. Intermittent sterilization method: the above two methods at atmospheric pressure, can only play a role in disinfection, and it is difficult to do Christine completely sterile. If the method of intermittent sterilization, you can kill all the microorganisms in the article. Specific method is: the items to be sterilized will be heated to 100 ° C, 15 ~ 30 minutes, to kill the nutrients. Then cool it and place it in a 37°C thermostat overnight to allow the residual spores to germinate into trophozoites. Repeat the above steps on the 2nd day, three times or so, to achieve the purpose of sterilization. This method does not require a pressurized sterilization pot, suitable for promotion, but the operation is troublesome and takes a long time.
d. Pressurized steam sterilization method: this is the fermentation industry, health care, food testing and microbiology laboratories in the most commonly used sterilization method. It is suitable for the sterilization of various heat-resistant, large-volume culture media, but also for the sterilization of glassware, work clothes and other items.
Pressurized steam sterilization is the items to be sterilized in an airtight pressurized steam sterilizer pot for a large amount of steam to make which the pressure rises. As the steam pressure rises, the boiling point of water also increases. When the steam pressure reaches 1.055 kg/cm2, the temperature inside the pressurized steam sterilizer can reach 121°C. In this case, microorganisms (including spores) are killed in 15 to 20 minutes, and sterilization is achieved. If the object of sterilization is sand, paraffin oil and other large area, containing more bacteria, poor heat transfer, the sterilization time should be extended appropriately.
In the pressurized steam sterilization, to draw attention to a problem is that before the constant pressure, be sure to exhaust the cold air in the sterilization pot, otherwise the steam pressure on the table and steam temperature does not have a corresponding relationship between, this will greatly reduce the sterilization effect.
3) Factors affecting sterilization
a. Different microorganisms or different ages of the same microorganisms have different sensitivities to high temperatures. Most of the microbial nutrients and viruses in 50 ~ 65 ° C, 10 minutes will be killed; but a variety of spores, especially spores are most resistant to heat, which is the strongest resistance to heat is thermophilic fat bacillus, to be in 121 ° C, 12 minutes before being killed. For the same microorganisms, young bacteria are more sensitive to heat than old bacteria.
b. The number of microorganisms obviously affects the effectiveness of sterilization; the greater the number, the longer the heat death time.
c. The ingredients and composition of the medium will also affect the sterilization effect. In general, the presence of proteins, sugars or fats improves heat resistance, pH near 7, the strongest heat resistance, biased toward the poles, the resistance to heat decreases, and different salts may have different effects on sterilization; solid media to be sterilized for a longer period of time than liquid media.
4) The effect of sterilization on the composition of the culture medium
a. pH generally decreases.
b.The production of turbidity or precipitation, which is mainly due to the chemical reaction of some ions to produce turbidity or ......
What is meant by sterilization? What kinds of sterilization methods are there Definition of sterilization:
Sterilization refers to the use of physical or chemical methods to kill all microorganisms, including pathogenic and non-pathogenic microorganisms as well as spores, so as to achieve the level of sterility assurance. After sterilization treatment, uncontaminated items are called sterile items. After sterilization, the area that is not contaminated is called the sterile area.
The resistance of microorganisms to sterilizing agents depends on the population density of the original presence, the strain, or the resistance conferred on the strain by the environment. Sterilization is necessary to obtain a pure culture and is a necessary technique in the food industry and medicine.
The process of killing or removing all trophic cells of microorganisms and their germ cells (or spores) from a culture medium, fermentation equipment, or other target, thereby achieving sterility.
Commonly used sterilization methods:
Heat sterilization
Heat sterilization uses high temperature to denature all proteins in the microbial cells, and enzyme activity disappears, resulting in cell death. Usually dry heat, wet heat and intermittent heating sterilization method.
Dry heat sterilization
Flame burning method or oven hot air sterilization method known as dry heat sterilization method of metal instruments or washed glassware into an electric oven, at 150 ~ 170 ℃ to maintain 1 ~ 2 hours, can be completely sterilized (including bacterial spores) purposes. Cauterization (incineration or burn) is one of the most thorough dry heat sterilization method, the scope of application is limited to the inoculation ring, inoculation needle sterilization or materials with pathogenic bacteria, animal carcasses burned, etc..
Moisture-heat sterilization by boiling water, steam and steam pressurization.
Pasteurization: because the earliest French microbiologist Pasteur used for fruit wine sterilization, so the name. This is a low-temperature sterilization method dedicated to milk, beer, fruit wine or soy sauce and other liquid flavored foods or seasonings that should not be sterilized at high temperatures.
Pasteurization is moist heat sterilization, and there are two ways to do this method [1], ① the classic lowtemperatureholdingmethod (LTH): processed for 30 minutes at 61.7 to 62.8°C; ② the more modern high temperature transient method (hightemperatureshorttime or Flushpoint (HTST): 15 min at 71.6°C or slightly higher. In the above methods, the best effect of steam pressurization, available atmospheric steam sterilization, but also in the high-pressure steam pot (generally use 1 kg / cm 2) sterilization, the steam temperature up to 121 ℃, can be heat-resistant spores in 30 minutes all killed. However, for certain substances that are easily destroyed by high pressure, such as some sugar or organic nitrogen-containing compounds, it is appropriate to sterilize at 0.6 kg/cm2 pressure (110 ℃) for 15 to 30 minutes.
Boiling disinfection method: the use of boiling at 100 ℃ for several minutes, generally used for the disinfection of drinking water.
Intermittent sterilization
Intermittent sterilization for three consecutive days, a daily steam sterilization method. This method is suitable for substances that cannot withstand temperatures above 100 ℃ and some sugar or protein substances. Generally in the normal atmospheric pressure with steam sterilization for 1 hour. Sterilization temperature does not exceed 100 ℃, does not cause the destruction of sugar and other substances, but can be intermittent culture during the germination of spores to kill, so as to achieve the purpose of complete sterilization.
Radiation sterilization
Radiation sterilization in certain conditions using the ray sterilization method. More commonly used are ultraviolet light, and other ionizing radiation (ray accelerated neutrons, etc.). Wavelength of 25000 ~ 80000 nanometers between the laser also has a strong ability to sterilize the wavelength of 26500 nanometers is most effective. Radiation sterilization method is limited to a certain material, due to the complexity of the equipment required, difficult to use widely.
Osmotic pressure sterilization
Osmotic pressure sterilization using high osmotic pressure solution sterilization method. In the high concentration of salt or sugar solution in the cell due to dehydration and the occurrence of plasma wall separation, can not carry out the normal metabolism, resulting in the death of microorganisms.
Chemical reagents sterilization
Most chemicals in low concentrations play a bacteriostatic role, high concentrations play a bactericidal role. Commonly used 5% carbolic acid, 70% ethanol and ethylene glycol. Chemical sterilization agent must have volatile, in order to remove the residual drug on the material after sterilization.
Chemical sterilization commonly used reagents are surface disinfectants, antimetabolites (sulfonamides, etc.), antibiotics, biopharmaceuticals antibiotics are a class of microorganisms or other organisms synthesized in the course of the life of the secondary metabolites or artificial derivatives, they are in a very low concentration can be inhibited or infected with it kind of organisms (including pathogens, viruses, cancer cells, etc.) of the life of the activity, and therefore can be used as an excellent It can be used as an excellent chemotherapeutic agent....
What are the common sterilization methods for medical devices? The verification method of medical device sterilization process is the compatibility with the bacterial process, the general sterilization methods include the following:
1, Moisture-heat sterilization:
Moisture-heat sterilization is the product is placed in the pressure cooker, the use of saturated steam under the pressure of the minimum temperature of 1210C, the heat and moisture is rapidly transferred to the sterilized product, the sterilization time of at least 15min. the latent heat of the steam is large, which can be The latent heat of steam is large, which can rapidly increase the temperature of the object, and the water molecules have strong penetration, which can easily coagulate and denature the proteins, therefore, wet heat sterilization is the most commonly used and more reliable sterilization method in heat sterilization. Medical devices suitable for wet heat sterilization include: clothes, sheets, PTFE, surgical instruments, silicone rubber, polypropylene, epoxy resin and so on. However, some polymers that are not resistant to high temperatures, such as polyurethane, cannot withstand such high temperatures and can only be sterilized by other methods.
2, dry heat sterilization:
Heat sterilization is to put the product in a hot air box, the use of dry air oxidation, kill all living microorganisms or eliminate pyrogenic methods. Dry heat sterilization is usually used at higher temperatures, ranging from 160 to 2500 ° C, depending on the temperature used, the exposure time can be up to 2 h. Dry heat sterilization conditions are generally more than 160 x 120min, 180. C x 60min or 250. C x 45min, other temperature and time parameters can also be used. In practice, the scope of dry heat sterilization is very limited, generally used in high-temperature-resistant glass appliances and metal surgical instruments, and some products are not only required to achieve the necessary level of sterility, but also the elimination of bacterial endotoxins (pyrogenic substances), the application of other sterilization methods is difficult to eliminate bacterial endotoxins, and dry heat sterilization temperature and time parameters are set to 250 ° C x 45min, you can remove the glass appliances of the pyrogenic substances.
3, ethylene oxide sterilization:
Packaging system of the material at least one with a certain degree of permeability, ethylene oxide sterilization is more commonly used in the field of medical equipment sterilization methods, ethylene oxide sterilization principle is through its relationship with the protein molecules on the sulfhydryl group (a SH), amino (a NH:), hydroxyl (a OH) and carboxyl (a COOH) and nucleic acid molecules on the imino (a NH), amino (a NH:), hydroxy (a OH) and carboxyl (a COOH). Alkylation reaction occurs in the imino group (NH:), hydroxyl group (OH) and carboxyl group (COOH), as well as on the nucleic acid molecule, resulting in the death of microorganisms, so as to achieve the sterilization effect. When ethylene oxide sterilization, the temperature, humidity, concentration of sterilizing gas and sterilizing time in the sterilizing cabinet are important parameters affecting the sterilization effect. Generally used sterilization conditions: temperature (55±10)oC, relative humidity (60±10)%, sterilization pressure 8xl05Pa sterilization time 120min. ethylene oxide is an alkylating agent, strong penetration, able to use a variety of packaging materials, at room temperature can kill a variety of microorganisms (including bacteria, spores, viruses, fungal spores, etc.) H. Applicable to biomedical polymer materials, such as natural rubber, polyethylene, polyethylene, polyethylene, polyethylene, polypropylene, polypropylene, polypropylene, polypropylene, polypropylene, polypropylene, polypropylene, polypropylene, polypropylene and other polymers. For example, natural rubber, polyethylene, polypropylene and polyvinyl chloride.
4, radiation sterilization:
Requirements for all packaging materials that constitute the packaging system can be resistant to irradiation ray treatment without aging and brittle;
Radiation sterilization is the sterilization of the product is placed in the appropriate radioactive source of radiation or the appropriate radiation source or the appropriate electron gas pedal generated by electron beams in the ionizing radiation generates free radicals by controlling the conditions of radiation to achieve the microbial killing method. The method of sterilizing is to control the radiation conditions to kill microorganisms. Sterilization of 1 ray is usually cobalt Co 60 or cesium Cs 137 as a radioactive source, the decay emits 1.33MeV and 1.17MeV two energy levels of rays, so that microbial DNA by the irrecoverable loss of the target people need to achieve the goal of 131. Radiation sterilization is usually used for surgical instruments, artificial prostheses, syringes, and sutures, and so on.
5, low-temperature plasma sterilization:
Low-temperature plasma sterilization is a new low-temperature sterilization technology developed in recent years. Plasma is a gas or steam affected by electric or magnetic fields, so that most of the molecules undergo ionization and formation. Plasma sterilizer is composed of power supply, excitation original, gas original, transmission system and sterilization chamber, after vacuuming through the water vapor, the steam is transformed into plasma under the action of electric field. Plasma sterilization advantages are reliable sterilization effect, low action temperature, sterilized instruments do not need to be placed in the air to remove residual gas, non-corrosive with. At present, many countries have begun to apply this technology, mainly for heat-resistant medical devices.
Summary:
The verification of the sterilization process is a necessary condition for sterility assurance, the sterility of the sterilized product depends on the selection of sterilization materials, suitable sterilization process in the production process, standardized GMP management quality management system, while the determination of the sterilization process should be taken into account in a comprehensive manner. ......
What are the sterilization methods in biologySterilization refers to the use of strong physical and chemical factors to kill all microorganisms inside and outside the object, including germs and spores.
Commonly used sterilization methods in the laboratory are: burning sterilization, dry heat sterilization, autoclaving sterilization.
(Sterilization and disinfection are different, with ultraviolet light treatment is disinfection.)
What are the commonly used sterilization methods in microbiology laboratories? The commonly used sterilization methods mainly include the following three: dry heat sterilization; humidification sterilization; ultraviolet sterilization
1. dry heat sterilization method
1.1 burning and flame sterilization: burning is mainly used for inoculation tool sterilization, burning on the flame can be achieved for the purpose of sterilization, flame sterilization is usually used for aseptic operation in the mouth of the test tube, Glass bottle mouth, silicone fluorine plastic plugs and other experimental items of sterilization, to prevent contamination of the mouth.
1.2 dry baking sterilization: the use of thermal radiation and dry hot air sterilization. Will be examined and sterilized items such as metal, glass, ceramic products after packaging, in the oven heated to 160 ℃, holding 2h can be completely sterilized. But should not exceed 170 ℃. In addition to cooling too quickly, sudden cold easy to cause glassware cracking. Dry heat sterilization when loaded into the dry oven items should not be tight, there should be a gap, conducive to the flow of hot air, too dense, resulting in uneven temperatures, part of the items sterilized incomplete.
2. Moisture-heat sterilization method
To increase the steam temperature by pressurizing, high-pressure steam sterilization, high temperature, sterilization effect is the best.
Precautions: 1. Completely exclude the cold air in the autoclave. The presence of cold air, in the same table under the pressure of the temperature value to be low, and cold air out of the less, the lower the temperature is more. In autoclaving, the cold air must be completely removed to ensure that the specified temperature is reached. Otherwise, although the pressure is reached, but the temperature does not meet the specified requirements, sterilization is not complete.
3. Ultraviolet light
Bactericidal spectrum is wide, but weak penetration, many factors affecting the bactericidal efficacy of certain restrictions.
Ultraviolet disinfection effect and ultraviolet intensity, irradiation time, temperature and humidity and other factors. The temperature of UV sterilization should be 20~40℃ and the relative humidity should be 40%~60%.
What are the commonly used disinfection and sterilization methods (1) natural disinfection: the use of daylight and other natural conditions to kill disease-causing microorganisms, to achieve the purpose of disinfection, known as natural disinfection.
①Sunlight exposure method: sunlight due to its heat, drying and ultraviolet effect, and has a certain bactericidal power. The size of the germicidal effect of sunlight by the region, season, time and other factors, the stronger the sunlight, the longer the irradiation time, the better the germicidal effect. Sunlight in the ultraviolet light through the atmosphere, due to heat dissipation and absorption and weakened, and can not all through the glass, therefore, must be directly exposed to the sun, in order to obtain the bactericidal effect. Sunlight exposure is often used to disinfect books, mattresses, bedding, blankets and clothing. Exposure should often be turned over the sun, so that all sides of the items can be in direct contact with sunlight, generally 4 to 6 hours of exposure to sunlight can achieve the purpose of disinfection.
② ventilation: ventilation can not kill microorganisms, but in a short period of time to make indoor and outdoor air exchange, reduce indoor pathogenic microorganisms. There are many ways to ventilate, such as using doors, windows or air windows, can also be used to ventilate the fan ventilation. The room should be ventilated regularly, ventilation time is generally not less than 30 minutes each time.
(2) physical sterilization method: the use of heat and other physical effects, so that the microbial protein and enzyme denaturation coagulation, in order to achieve the purpose of disinfection, sterilization, known as physical sterilization method.
① Combustion method: a simple and easy, rapid and thorough and effective sterilization method, but the destructive nature of the items, mostly used for high heat resistance, or has been with pathogenic bacteria and no value of the items retained, such as paper, dressings contaminated with certain bacteria or viruses, enameled items such as the bidet, you can also use the flame combustion disinfection sterilization, should be the first basin washed and dried, and then poured a small amount of 90% alcohol, ignited! Slowly turn the tub, so that its inner surface is completely burned by the flame. Application of this method, pay attention to safety, must be away from flammable or explosive substances to avoid causing fire.
② boiling method: is an economical and convenient sterilization method, generally wait for the water to boil after timing, boiling 10-15 minutes can kill germ-free bacteria. Can be used for food utensils, towels, handkerchiefs and other items that are not afraid of humidity but resistant to high temperature sterilization.
3 high-pressure steam sterilization method: the use of high pressure in the autoclave and high heat release potential for sterilization, this method is a strong bactericidal force, is the most effective physical sterilization method. To be steam on the autoclave, add the valve and then steam for 15 minutes, suitable for sterilization of cotton, dressings and other items.
(3) chemical disinfection sterilization method: chemical disinfection sterilization method is the use of chemical drugs to penetrate the bacterial body, destroy its physiological function, inhibit bacterial metabolic growth, thus playing a disinfection role. Family commonly used chemical disinfection sterilization methods are the following three.
①Wiping method: wipe the surface of the contaminated object with chemical solution, commonly used in the ground, furniture, display items disinfection. Such as 0.5% -3% bleach clarifying solution, 84 disinfectant and other chlorine disinfectant (now sold in the market, to look at the expiration date and use), wipe walls, beds, tables and chairs on the ground and toilets.
② immersion method: the items to be disinfected immersed in disinfectant, often used in the living utensils can not or inconvenient to steam. The length of the soaking time varies depending on the nature of the item and the solution. Such as 1% -3% bleach clarifying solution soak tableware, toilet needs 1 hour, with 0.5% "84" disinfectant soak needs 15 minutes, and with 0.02% high efficiency disinfectant tablets soak only 5 minutes, you can achieve the purpose. If the soaking of vomit and excreta, not only the concentration of disinfectant to be doubled, and the soaking time should be doubled.
3 ③ fumigation method: is the use of disinfectant drugs produced by the gas to disinfect. Commonly used in infectious patients have lived in the room air and indoor surface disinfection.
Formalin (formaldehyde) + potassium permanganate: 25-40 ml of formalin per cubic meter, 15-30 grams of potassium permanganate, the two medicines are placed together that is to produce gas, can achieve the purpose of disinfection. When disinfecting, the doors and windows must be closed tightly for 12-24 hours, and then open the doors and windows for ventilation after disinfecting, this method is effective for all kinds of infectious diseases caused by bacteria and viruses.
Vinegar: 3-10 ml of vinegar per cubic meter, add water 2-3 times heated fumigation for indoor air disinfection, effective for the prevention of influenza and other respiratory infectious diseases.