NAME: Gabinete, Nathaniel T.

DATE: October 07, 2022

YEAR/SECTION: BS Bio 2A

Laboratory Activity 4
Preparation of Culture Media

Introduction

In the field of microbiology, microbial growth is always observed and experimented

inside laboratories. Specific combinations of nutrients and other materials are referred to as
culture media or growth media, and they are used to promote the development of
microorganisms like bacteria and fungi. According to Mettler Toledo (2022), every
microbiology laboratory regularly prepares culture media formulations such as liquid
growth media and culture media based on nutrient agar. The creation of culture media is a
multi-step process that requires caution to prevent cross-contamination. For the proper
promotion of microbiological development, culture medium preparation must be done
precisely.

When preparing culture media, it is important to keep track of the surroundings

always. To prevent the formation of molds and cross-contamination, utilizing aseptic
technique is necessary. A set of techniques and procedures called aseptic techniques help
protect against harmful bacteria. Several microorganisms are found everywhere, so using
aseptic can help prevent laboratory equipment from being infested by bacteria that may
affect the agar.

Objectives

1. To demonstrate agar pouring correctly; and

2. To apply aseptic technique while doing agar pouring.

Methodology

In preparing culture media, there are a lot of processes involved. First is wrapping a
petri dish with paper in order to prevent contamination from outside. Put the petri dish in
the center of the paper then fold each side of the paper making sure that the petri dish is fully
covered. Fold the excess paper in between by overlapping them at the center. Fold the sides
of the paper below the dish and make sure to keep the circular shape of the petri dish by
pressing the edges.
 Next is creating a cotton plug for the Erlenmeyer flask using a roll of cotton, gauze,
and thread. Grab a roll of cotton then fold each side making sure that the ends meet the
center. Roll the cotton horizontally. Make sure that it is tightly rolled and that there are no
air spaces. Wrap the rolled cotton with the gauze and secure it using a thread.

Create the agar mixture after. Prepare 500 mL of water and put ½ tbsp of gelatin
powder (the amount depends on the mL of water used), a pinch of salt, Vitamin C, and beef
broth. Make sure to mix properly to dissolve every solute thoroughly. Once done, cook the
mixture under medium heat for 15-20 minutes. Carefully transfer the mixture into the
Erlenmeyer flask once finished. Cover the flask using a cotton plug and let it sit for several
minutes to cool down at room temperature.

Before using any laboratory equipment, it is essential to sterilize every piece of

equipment to be used by steaming it for 15-20 minutes under medium to high heat. Disinfect
the work area too by spreading 70% solution alcohol on the surface and wiping it using a
strip of clean tissue. After that, wear proper laboratory attire (lab gown, sterile gloves, and
bouffant cap) before performing agar pouring.

Lastly, perform agar pouring. Prepare the mixture using the sterile petri dish and the
Erlenmeyer flask. Carefully unwrap the petri dish and remove the plug of the Erlenmeyer
flask before pouring. The alcohol lamp should be lit. Twist the flask once and place the mouth
over the flame. Use the opposite hand to perform the same action with the petri dish, paying
attention to how the fingers should be positioned. Then, fill the petri dish with the mixture
until it is halfway full. Be sure to carry out this action close to the flame. The flask's mouth
should then be heated under a flame to disinfect it before being quickly sealed with a cotton
plug. Any leftover mixture can be stored inside the refrigerator for future use.

Before storing the petri dish, make sure to let the agar harden for a few hours at room
temperature. Once solidified, wrap it again with the paper used and store it inside the
refrigerator for 24 hours before using.
Results and Discussion

Figure 1. Wrapping of Petri Dish

Figure 2. Agar Pouring with Aseptic Technique

 Figure 3. Culture Media (Hardened)

After almost 3 hours, the agar solidified as seen in Figure 3. After 24 hours of being
stored inside the refrigerator, no molds have formed on the surface of the culture media.

Conclusion

In conclusion, good agar pouring may be accomplished with the right processes and
instructions. If done correctly, there shouldn't be any issues like mold growth or agar that
doesn't solidify after several hours. To avoid unwanted contamination of the created culture
media, it is also crucial to use disinfection procedures, such as the aseptic technique, in which
the equipment is heated by a flame before and after pouring.
References

Mettler Toledo (2022). Culture Media Preparation. Mettler-Toledo International Inc. All
Rights Reserved. Retrieved from https://www.mt.com/in/en/home/applications/
Laboratory_weighing/Culture-Media-Preparation.html