Methods of Cleaning And Sterilization in the Laboratory

Microbiology, tissue culture, medical, equipment manufacturing labs, and many research labs and industries need strict sterile environments for their diverse operations. Experiments, specifically those involving cell lines or microorganisms need to be conducted in a controlled environment. Contamination not only voids experiments, but also wastes effort, time, and money and when involving patients, it poses serious health risks. It is essential to be well-versed in laboratory sterilization techniques.

Sterilization is a process of killing bacteria, fungi, viruses, and other microorganisms.[1] Sterilization of labs and equipment prevents the transfer of unwanted microbes to lab cultures or infects patients during medical procedures.[1]

This article addresses why sterilization is essential, some of the commonly used methods in labs for sterilization of surfaces and equipment, and factors to consider while choosing a sterilization method.

In the lab or medical space, aseptic conditions are required for the following [2]:

Bringing down the pathogenic load on surfaces

Eliminating microbes’ replication sites

Preventing corrosion and contamination of high precision and expensive tools with intricate parts

Reducing biological changes in model organisms

Ensuring safe use of invasive and non-invasive medical devices[2]

Protecting cultures and subjects from getting contaminated and infected

Providing aseptic environments within the lab allows for accurate and precise experimentation on living cells and tissues without undesired parameters to change the physiological and metabolic processes of any subjects involved.[2]

Today, different types of sterilization methods are used in labs based on the equipment or material to be sterilized. Though the result of all the techniques is approximately similar, they are different in their approach. 

Wet Heat or Moist Heat Sterilization

It’s one of the most popular techniques used in labs for the sterilization of equipment and glassware.[3] It involves autoclaving: heating material in moist steam at 121°C and 15 psi for 15-20 minutes to effectively sterilize. It kills and inactivates viruses, bacteria, spores, and other microbes by hydrolysis and coagulation of cellular proteins.[3]

Another wet heat technique is “boiling”.[4] The sterilization through boiling is carried out in a water bath at 100℃ for 30 minutes. It’s mainly used to sterilize rubber goods, syringes, and surgical instruments.[4]

This technique is economical, reliable, and non-toxic. Further, it has more penetrating power than the dry heat approach. However, caution is required while using the autoclave equipment to avoid burns.

Dry Heat Sterilization

This process does not involve water or steam to sterilize equipment.[3] Thus, hydrolysis of microbes’ proteins can’t take place. This is one notable difference between dry heat and wet heat sterilization.[3] 

Dry heat involves flaming, hot air ovens, and incineration, killing and inactivating microbes by oxidizing their cellular components. To achieve this, the temperature is raised to 325 degrees Fahrenheit or higher under normal air pressure for efficient sterilization.[5]

Incineration is mainly used to sterilize inoculation loops; flaming has uses in sterilizing scalpels, needles, and scissors; whereas a hot air oven is used to sterilize glassware, some metal devices, and dry materials like powders.

The advantages of heat sterilization are that it’s simple, cost-efficient, easy to install, and an excellent approach for steam-sensitive.[6]

Tyndallization

It’s also known as fractional sterilization.[4] The process is performed at 100°C in a steam sterilizer for 20 minutes, followed by overnight incubation at 37°C. The cycle is repeated for two successive days. It’s an effective technique to kill and inactivate microbes and heat-resistant endospores.[4]

However, the technique is not completely reliable, as some spores might survive the process and activate.[7] Thus, it’s only used when the materials need to be sterilized and can’t stand pressurized heating. For example, it is commonly used to sterilize heat labile media containing milk, sugar, and gelatin.[7]

Autoclaving

The method uses high-pressure steam. It’s performed at 121°C (or 250 ℉) and 15 lbs per square inch of atmospheric pressure for 15-20 minutes.[8] It’s an effective method to kill or inactivate spores, fungi, bacteria, and viruses. It’s the most commonly used technique in labs because it’s cheap and economical, provides results in a short time, requires no additional tools or chemicals, and offers effective penetration in objects for their effective sterilization.[8]

Pasteurization

Pasteurization is a type of moist heat sterilization but it operates at a temperature of less than 100°C.[4] It is generally used to eliminate bacteria and pathogens from packaged and non-packaged foods (such as fruit juices and milk). A phase change of the product can be inhibited by the heat treatment and cooling process. However, it doesn’t eliminate contaminants, such as spores.[4]

It kills and inactivates microbes by coagulation and denaturation of proteins, breaking DNA strands, and destroying cell membranes. 

Filtration

Filtration is one of the quickest sterilization techniques, which does not require heat.[5] Through this process, microbes are blocked from passing through filters with too small pores. Thus, microbes are not killed but separated from a solution or media. Membrane filters used in the process are made of cellulose esters and have a pore diameter of 0.2 μm.[5] 

However, this method is not suitable for viral removal due to its size being smaller than the filter pores and can easily pass through.[5]

Chemicals

Heating some equipment or tools can often damage them.[9] Thus, liquid chemicals and gasses are the go-to approaches to sterilizing such delicate materials. Gasses provide better penetration, achieving effective sterilization. Ethylene oxide and carbon dioxide are the most commonly used gasses for the sterilization process. Ozone gas is used to sterilize organic matter.[9] 

Disinfecting agents like quatricide and super oxidized or super hypochlorous water are used to clean behavioral testing equipment. Seventy percent (70%) of ethanol has application in sterilizing lab equipment.[10] 

Radiation

Radiation is one of the most effective sterilization techniques used in labs.[9] It utilizes electromagnetic radiation X-rays, Gamma-rays, and Ultraviolet rays to pulverize the DNA of microbes and kill them.[9] However, each ray differs in its effectiveness and penetration ability. For example, UV rays have low penetration ability and less effectiveness. For large-scale sterilization, gamma rays, and X-rays have more extensive applications. However, radiation is hazardous to human health and requires extra safety precautions.[9]

Sterilizing biological safety cabinets between uses is routinely achieved by UV irradiation. A large package or pallet load of medical devices can be sterilized with X-rays. Syringes, cannulas and IV sets, needles, as well as food, are commonly sterilized using gamma radiation.[9]

Sterilizing techniques are essential for labs and industries to eliminate and reduce bacteria, viruses, fungi, and spores to achieve desired results without the risk of contamination. Today, various sterilization methods are available to sterilize equipment and materials. It includes wet heat sterilization, dry heat sterilization, pasteurization, radiation, filtration, and chemical-based.

However, not every technique can be used to sterilize every piece of equipment or material. Consider factors such as purpose, ease of method, the material composition of the equipment or tool being sterilized, and budget before choosing a sterilizing technique suitable for your lab needs.

Check out this high-tech, safe, and automated Research Autoclave and Cellulose Acetate Membranes for your lab sterilization processes, such as equipment, glassware, plasticware, and chemicals.

1. Importance of Sterilization (2020).   
2. Importance of Instrument Disinfection & Sterilization.
3. Oswald Nick (2021). 6 Laboratory Sterilization Methods.
4. Kumar Abhay, Murthy Narasimha L., Jeyakumari A., and Laly S. J. Sterilization Techniques Used in Microbiology. 
5. 5 Common Methods of Lab Sterilization (2021). 
6. Cephas Casey. Choosing The Best Sterilization Method For Your Product.
7. Tynadallization. 
8. What is autoclaving and how is it relevant to liquid handling instruments? 
9. Different sterilization methods were used in the laboratory (2018).
10. The Mouse Nose Knows: Cleaning The Behavioral Testing Apparatus (2017).