Week 9
Posted by drstocksblog on October 23, 2009
Exercise 12: Determining the Number of Bacteria in Food
For this exercise you will work in a team to determine the number of bacteria per gram of hamburger. Half of the teams will have meat that has been left out at room temperature for a number of hours and the other half will have meat that has been continuously refrigerated. We will compare the results of the two teams for the return lab.
We are using the pour plate technique to mix a known portion of the food with agar, letting it incubate, and counting the number of resulting colonies. This will give you an estimate of the number of bacteria in a gram of the food.
Dilution
- Mixing a gram of hamburger with agar will result in a huge number of colonies — clearly more than you can count. A plate with more than 300 colonies is considered (and reported as) Too Numerous to Count (or TNTC).
- Note that a plate with less than 30 colonies is considered Too Few to Count (TFTC).
- Sooo the hamburger must be diluted and then the dilution is placed in the agar.
- You are going to doing three dilutions with the hamburger:
- 20 in 180 or 20/200 = 1 to 10 (or 1/10 or 1:10 or 0.1)
- then that will be diluted 1 in 99 = 1 to 100 (or 1/100 or 1:100 or 0.01) [total dilution 1/10 and 1/100 = 1/1,000]
- and that will be diluted 1 in 99 = 1 to 100 (or 1/100 or 1:100 or 0.01) [total dilution 1/1000 and 1/100 = 1/100,000]
- We’ll go over this at the start of lab.
Pour Plate
- You will be placing known quantities of your dilutions in empty sterile petri dishes.
- We are duplicating everything this week. [Why is replication necessary and important?]
- Then mixing sterile melted agar with your dilution in the plate, allowing it to solidify, and then incubating it at 37 degrees C for 48 hours.
Determining the Number of Bacteria:
- For the return lab you will count the number of colonies on those plates that can be accurately counted:
- those containing between 30 and 300 colonies.
- Then you will calculate the number of bacteria per gram of meat by the following formula:
- # bacteria/gram = # colonies x 1/mL plated x 1/dilution plated
COME TO LAB PREPARED BY CAREFULLY READING THE LAB AND THIS BRIEFING!!!
There is NO Lab Quiz this week. HOWEVER — there is a set of questions on dilutions that is due on your return lab on Wednesday or Thursday!!! NO LATE ASSIGNMENTS WILL BE ACCEPTED WITHOUT A DOCTOR’S EXCUSE!!!!! It is posted in Vancko Hall.
