Name:
Date:
Aseptic Technique and Streak Plate Technique
Materials:
 Bunsen burner
 Test tube rack
 Permanent marker
 Culture tube containing bacteria
 3 sterilized culture tubes
 Petri dish with agar
 Wire inoculating loop
Aseptic Technique
Procedure:
1. To inoculate a culture tube, hold the loop like a pencil in your dominant hand
and hold the culture tube in your non-dominant hand. Remove the cap from
the tube with your dominant hand, holding it there. Flame the mouth of the
tube by passing it through the flame at a 45-degree angle.
2. Inoculate a sterile culture tube with the unsterilized loop. Label the tube and
place it in the test tube rack.
3. To sterilize the loop, light the Bunsen burner and adjust the flame until there
is a center blue cone. Flame the loop by holding the loop at a 45-degree angle
with the end where the wire meets the handle in the flame. Pull the wire
through slowly allowing it to burn red-hot until the loop has also been
through the flame. Allow the loop to cool for 30 seconds.
4. Inoculate a sterile culture tube with the sterilized loop with the same
technique as step 1. Label the tube and place it in the test tube rack.
5. Re-flame the loop. Hold both the bacteria culture tube and the un-inoculated
tube in your non-dominant hand. Remove the caps from the tubes with your
dominant hand, holding them there. Flame the mouths of the tubes by
passing them through the flame at a 45-degree angle.
6. Use the inoculating tube to remove a small amount of the bacteria from the
tube and transfer it to the un-inoculated tube by placing the loop at the
bottom of the agar and using a zigzag motion, pull the loop up to the top of
the agar.
7. Re-flame the loop before putting it down.
8. Re-flame the mouth of the tubes and replace the caps. Label the inoculated
tube and place it in the test tube rack.
9. Place the test tube rack in the inoculating chamber overnight.
Streak Plate Technique
Procedure:
1. Divide the plate into quadrants by drawing lines across the cover of the agar
plate. Label them 1, 2, 3, 4.
2. Flame the loop. Using aseptic technique, remove a small amount of bacteria
from the bacteria culture tube. Replace the culture tube in the test tube rack.
3. Open the agar plate by holding the cover at a 45-degree angle over the plate.
Zigzag the loop across the surface of the agar in quadrant 1 only. Replace the
cover and flame the loop.
4. Make a quarter-turn of the plate. Open the cover. Drag the loop from
quadrant 1 to quadrant 2 and zigzag the loop across the surface of the agar in
quadrant 2 only. Replace the cover and flame the loop.
5. Repeat step 4 for quadrant 3 and 4.
6. Flame the loop before putting it down. Place the inoculated agar plate in the
incubation chamber overnight.
Observation and Discussion Questions
1. Examine the three culture tubes you prepared. Compare the results of each
tube to each other and provide an explanation for what you find. Sketch your
observations.
2. How does the culture tube that was inoculated with the unsterilized loop
demonstrate the importance of not putting the loop down while you work
and re-flaming it so often throughout the procedure?
3. What would happen if you put the caps of the tubes down while you were
inoculating the tubes and then recapped the tubes with them? Why is it also
important to flame the mouths of the tubes?
4. Examine the culture plate that you prepared. Describe and compare what
you find in each quadrant. Sketch your observations below.
5. What would happen if you completely removed the cover of the Petri dish
while you were streaking the plate?
6. How does what you find in quadrant 4 demonstrate the importance of using
the streak plate technique?
7. How would you use the streak plate technique to study a sample of bacteria
taken from the surface on which you are working?