Different sterilization methods used in the laboratory

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Sterilization can be achieved by a combination of heat, chemicals, irradiation, high pressure and filtration like steam under pressure, dry heat, ultraviolet radiation, gas vapor sterilants, chlorine dioxide gas etc. Effective sterilization techniques are essential for working in a lab and negligence of this could lead to severe consequences, it could even cost a life.

So what are the most commonly used methods of sterilization in the laboratory, and how do they work? Read on…

Heat Method: This is the most common method of sterilization. The heat is used to kill the microbes in the substance. The extent of sterilization is affected by the temperature of the heat and duration of heating. On the basis of type of heat used, heat methods are categorized into-

(i) Wet Heat/Steam Sterilization- In most labs, this is a widely used method which is done in autoclaves.. Autoclaves use steam heated to 121–134 °C under pressure. This is a very effective method that kills/deactivates all microbes, bacterial spores and viruses. Autoclaving kills microbes by hydrolysis and coagulation of cellular proteins, which is efficiently achieved by intense heat in the presence of water. The intense heat comes from the steam. Pressurized steam has a high latent heat and at 100°C it holds 7 times more heat than water at the same temperature. In general, Autoclaves can be compared with a typical pressure cooker used for cooking except in the trait that almost all the air is removed from the autoclave before the heating process starts. Wet heat sterilization techniques also include boiling and pasteurization.

(ii) Dry heat sterilization- In this method, specimens containing bacteria are exposed to high temperatures either by flaming, incineration or a hot air oven. Flaming is used for metallic devices like needles, scalpels, scissors, etc. Incineration is used especially for inoculating loops used in microbe cultures. The metallic end of the loop is heated to red hot on the flame. The hot air oven is suitable for dry material like powders, some metal devices, glassware, etc.

Filtration is the quickest way to sterilize solutions without heating. This method involves filtering with a pore size that is too small for microbes to pass through. Generally filters with a pore diameter of 0.2 um are used for the removal of bacteria. Membrane filters are more commonly used filters over sintered or seitz or candle filters. It may be noted that viruses and phage are much smaller than bacteria, so the filtration method is not applicable if these are the prime concern.

Radiation sterilization: This method involves exposing the packed materials to radiation (UV, X-rays, gamma rays) for sterilization. The main difference between different radiation types is their penetration and hence their effectiveness. UV rays have low penetration and thus are less effective, but it is relatively safe and can be used for small area sterilization. X-rays and gamma rays have far more penetrating power and thus are more effective for sterilization on a large scale. It is, however,  more dangerous and thus needs special attention. UV irradiation is routinely used to sterilize the interiors of biological safety cabinets between uses. X-rays are used for sterilizing large packages and pallet loads of medical devices. Gamma radiation is commonly used for sterilization of disposable medical equipment, such as syringes, needles, cannulas and IV sets, and food.

Chemical method of sterilization: Heating provides a reliable way to get rid of all microbes, but it is not always appropriate as it can damage the material to be sterilized. In that case, chemical methods for sterilization is used which involves the use of harmful liquids and toxic gases without affecting the material. Sterilization is effective using gases because they penetrate quickly into the material like steam. There are a few risks, and the chances of explosion and cost factors are to be considered.

The commonly used gases for sterilization are a combination of ethylene oxide and carbon-dioxide. Here Carbon dioxide is added to minimize the chances of an explosion. Ozone gas is another option which oxidize most organic matter. Hydrogen peroxide, Nitrogen dioxide, Glutaraldehyde and formaldehyde solutions, Phthalaldehyde, and Peracetic acid are other examples of chemicals used for sterilization. Ethanol and IPA are good at killing microbial cells, but they have no effect on spores.


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