Plate Exposure Method: Principle and Procedure

Explore the Plate Exposure Method, a key technique in environmental and industrial microbiology. Understand its principle, applications in various fields, and the benefits and drawbacks of this cost-effective, simple procedure for monitoring airborne microbial contamination

Principle

The principle of the plate exposure method, also known as the settle plate method or passive air sampling, lies in the physical settling of airborne particles onto the surface of a nutrient agar in a Petri dish. As air moves over the plate, microorganisms in the air settle onto the agar surface. Once on the agar, microorganisms find nutrients and moisture that allow them to grow and multiply, forming visible colonies.

SOP for Plate Exposure Method

Preparation of Agar Plates

Begin with preparation of agar plates. Use the appropriate type of agar medium for the microorganisms you expect to encounter. For instance, nutrient agar is a good general-purpose medium. If you’re focusing on bacteria, you might choose tryptic soy agar, and if you’re focusing on fungi, you might choose Sabouraud agar.
Weigh the correct quantity of the chosen agar medium (usually indicated on the product’s label, typically around 15-20g for 500ml distilled water).
Dissolve the agar medium in distilled water by heating in a suitable container such as an autoclavable flask or bottle. Use a hot plate or autoclave (Working Principle of Autoclave) for heating if necessary.
Autoclave the solution to sterilize it, typically at 121 degrees Celsius for 15 minutes.
Allow the sterilized agar to cool but not solidify (around 45-50 degrees Celsius) and then pour it into sterile Petri dishes. Cover the dishes immediately after pouring to prevent airborne contamination.
Let the agar solidify completely at room temperature.
Store the prepared agar plates in a refrigerator until they are ready for use.

Plate Exposure

Take the prepared agar plates to the area where you want to collect microbial samples.
Remove the lid of the Petri dish and expose the agar surface to the air for a predetermined amount of time (often between 5 to 15 minutes depending on the study design).
Ensure that the agar plate is placed on a flat, stable surface during exposure, and avoid moving or shaking the plate.
Make sure that you don’t touch the inside of the lid or the agar surface to avoid contamination.
After exposure, immediately replace the lid of the Petri dish.

Incubation

After exposure, the plates should be incubated to allow the microorganisms to grow.
Seal the Petri dishes with parafilm or laboratory film to prevent contamination during incubation.
Incubate the plates upside down to prevent condensation from dripping onto the agar surface. The side of the Petri dish containing the agar should face upwards.
Incubate at a temperature suitable for the growth of the expected microorganisms, commonly at 37 degrees Celsius for bacteria and 25-30 degrees Celsius for fungi.
Allow the plates to incubate for a suitable period, typically 24-48 hours for bacteria and up to 5-7 days for fungi.

Colony Counting

After incubation, microbial colonies should be visible on the agar surface.
Count the number of colonies on each plate. Each colony represents a CFU (Colony Forming Unit), which is generally assumed to have originated from a single microorganism.
Record the number of CFUs, the incubation time, the exposure time, and any other relevant details for each plate.

Identification and Analysis

If necessary, you can proceed with identification of the microorganisms based on their morphology (color, size, shape, texture, etc.), gram staining and/or other biochemical tests.
Be sure to follow the correct procedures for the specific identification methods you are using.
Always handle cultures (Disinfectant validation protocol) with appropriate biosafety precautions.

Uses- Plate Exposure Method

Environmental Monitoring: The method is often used to measure the level of microbial contamination in the air within specific environments such as hospitals, schools, offices, and industrial settings (e.g., food processing plants, pharmaceutical manufacturing facilities). It is useful in assessing the effectiveness of sanitation procedures, the functioning of air filtration systems, and overall indoor air quality.
Food and Pharmaceutical Industry: It is used to ensure sterility in critical areas and to evaluate the effectiveness of cleaning and disinfection procedures. It’s crucial in maintaining quality control and adhering to regulatory requirements.
Research Studies: The method can be used in research studies investigating the diversity and distribution of airborne microorganisms.

Advantages- Plate Exposure Method

Simplicity and Cost-effectiveness: The procedure is relatively straightforward and does not require specialized equipment, making it cost-effective.
Visibility of Results: The method allows for easy observation of results as microbial colonies can be visually counted.
Potential for Further Analysis: Colonies grown on plates can be further analyzed to identify the species of microorganisms present.

Limitations- Plate Exposure Method

Not Quantitatively Accurate: The method only captures a fraction of the airborne particles – those that happen to settle onto the plate during the exposure period. Thus, it does not provide a complete or quantitatively accurate picture of airborne microbial contamination.
Depends on Environmental Conditions: The efficiency of this method can be affected by environmental conditions such as air currents, temperature, and humidity.
Time Consuming: The method requires an incubation period to allow microbial growth, which can take several days.
Requires Sterile Techniques: There is a risk of contamination during plate preparation and handling. Proper sterile techniques are needed to ensure that the only microorganisms counted are those from the air sample.

Uses

Environmental Monitoring: The method is often used to measure the level of microbial contamination in the air within specific environments such as hospitals, schools, offices, and industrial settings (e.g., food processing plants, pharmaceutical manufacturing facilities). It is useful in assessing the effectiveness of sanitation procedures, the functioning of air filtration systems, and overall indoor air quality.
Food and Pharmaceutical Industry: It is used to ensure sterility in critical areas and to evaluate the effectiveness of cleaning and disinfection procedures. It’s crucial in maintaining quality control and adhering to regulatory requirements.
Research Studies: The method can be used in research studies investigating the diversity and distribution of airborne microorganisms.

Advantages

Simplicity and Cost-effectiveness: The procedure is relatively straightforward and does not require specialized equipment, making it cost-effective.
Visibility of Results: The method allows for easy observation of results as microbial colonies can be visually counted.
Potential for Further Analysis: Colonies grown on plates can be further analyzed to identify the species of microorganisms present.

Limitations

Not Quantitatively Accurate: The method only captures a fraction of the airborne particles – those that happen to settle onto the plate during the exposure period. Thus, it does not provide a complete or quantitatively accurate picture of airborne microbial contamination.
Depends on Environmental Conditions: The efficiency of this method can be affected by environmental conditions such as air currents, temperature, and humidity.
Time Consuming: The method requires an incubation period to allow microbial growth, which can take several days.
Requires Sterile Techniques: There is a risk of contamination during plate preparation and handling. Proper sterile techniques are needed to ensure that the only microorganisms counted are those from the air sample.