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Design Project

The document is a set of instructions for students in a microbiology lab course on how to perform serial dilutions and plating of a bacterial culture. It includes an introduction explaining why dilution is necessary to get an accurate colony count. The materials, safety tips, and 22 step procedure are then outlined. Students will already know proper aseptic techniques. The goal is to end up with 6 labeled petri dishes containing diluted samples that will be incubated for 24 hours to allow colony growth before counting to determine CFU (colony forming units).

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70 views

Design Project

The document is a set of instructions for students in a microbiology lab course on how to perform serial dilutions and plating of a bacterial culture. It includes an introduction explaining why dilution is necessary to get an accurate colony count. The materials, safety tips, and 22 step procedure are then outlined. Students will already know proper aseptic techniques. The goal is to end up with 6 labeled petri dishes containing diluted samples that will be incubated for 24 hours to allow colony growth before counting to determine CFU (colony forming units).

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Copyright
© © All Rights Reserved
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You are on page 1/ 5

Sariah VanderVeur

ENGL 2100 Section 403 - Jorgenson

Time 10:00 a.m.

Design Project
Design Project Letter of Transmittal

April 29, 2020

ENGL 2100 Class


SLCC Taylorsville Campus
4600 S Redwood Rd
Salt Lake City, UT 84123

Subject: ENGL 2100 Design Project – Sariah VanderVeur

To whom it may concern,

Regarding the Design Project assignment for ENGL 2100, Spring Semester 2020, I am submitting
this letter of transmittal as requested to notify you of its timely delivery.

The following document is a set of instructions for the serial dilution and plating of a bacterial
culture for a college microbiology lab course. Students will already know proper aseptic,
sterilization, and techniques needed to follow the instructions. Included in the instructions are
an introduction, materials needed, safety reminders and tips, and the procedure.

Let me know if you have any questions, otherwise I’ll look forward to receiving a perfect grade.

Best Regards,

Sariah VanderVeur
Sariah VanderVeur, Student
Salt Lake Community College
[email protected]

2
Serial Dilutions and Plating

Introduction
Determining the number of bacteria in a sample can be difficult due to the high density in a
sample. A solution to this issue is counting colonies on a petri dish to determine the number of
colony forming units, or CFU. Although, even trying to count colonies can be difficult when
there are too many on the petri dish. In order to get an accurate colony count dilution of the
sample is necessary so that each colony can be distinguished. An accurate CFU count lies from
30-300 and can be determined by the following formula:

𝐶𝐹𝑈 # 𝑜𝑓 𝑐𝑜𝑙𝑜𝑛𝑖𝑒𝑠 𝑝𝑙𝑎𝑡𝑒𝑑 ∗ 𝑑𝑖𝑙𝑙𝑢𝑡𝑖𝑜𝑛 𝑓𝑎𝑐𝑡𝑜𝑟


=
𝑚𝐿 𝑣𝑜𝑙𝑢𝑚𝑒 𝑝𝑙𝑎𝑡𝑒𝑑

Materials Needed
• 10 mL source culture • Sterilized agar
• 5 test tubes • 95% ethanol
• 0.85% saline solution • 6 empty petri dishes
• Micro-pipette • Parafilm
• Bunsen burner • Click counter

Important Safety Reminders and Tips


• Tie long hair back
• Keep gloves, lab coat, and goggles on at all times
• No food or drink in the lab
• Do not set sterilized equipment on the counter, use it as fast as possible to reduce air-
borne contamination, and replace in ethanol when done
• Always keep Petri dish lid on, or at a 45° angle while plating samples
• Keep agar solution in a 45 °C water-bath while not in use

Dilution and Plating


1. Label the source culture tube as 0 and the 5 test tubes 1-5
2. Label 6 empty petri dishes 0-5
3. Place 9 ml saline solution in test tubes 1-6

Steps 1-3

1
4. Loosen the lids of tubes 0 and 1 and Step 5 Step 6
flame the edges of the tubes before
transferring the sampl
5. With a sterilized micro-pipette tip
transfer 1 ml of the source culture,
tube 0, into the petri dish labeled 0
6. Transfer another 1 ml of sample from tube 0 into tube 1
7. Flame the edges of tube 1 before closing then shake gently to mix contents, dispose of
micro-pipette tip
8. With a new micropipette tip repeat the process with the next tube and plate
9. Flame the edges of tubes 1 and 2 then transfer 1 ml of solution from tube 1 into petri
dish 1 and another 1 mL into tube 2.
10. Flame the edges of tube 2 and gently shake to mix, dispose of micro-pipette tip.
11. Repeat steps 9 and 10 starting with tube 2 then going down the chain as shown below

2 3 4 5

Step 11
12. Once all the samples have been diluted and plated it will be time to add agar solution
13. Remove the sterilized agar kept from the 45 °C water bath, dry it off, and flame the mouth
and neck
14. WORK QUICKLY AND ASEPTICALLY
15. Holding the petri dish slightly open add agar to the dilution liquid until it covers about 2/3
of the area

Step 12
16. Mix the agar and dilution liquid by gently swirling the dish
17. Flame the mouth of the agar solution again before moving onto the next dish
18. Follow steps 15-17 for the remaining petri dishes
19. Once petri dishes 0-6 all have agar solution set them aside and leave them undisturbed for 10
minutes
20. Seal the petri dishes with parafilm and invert them
21. Incubate dishes at 35 °C for 24 hours
22. Once incubated use cell counter to assist you in counting the number of colonies per plate,
then determine the number of CFU

2
Bibliography

Steane, Richard. “DILUTION PLATING.” Microbiological Techniques - Dilution Plating,


www.biotopics.co.uk/microbes/tech3.html.

“11 Pour Plate Method Best Practices.” Microbiologics Blog, 8 Mar. 2018, blog.microbiologics.com/11-pour-plate-
method-best-practices/.

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