BISC209: Aseptic Transfer

Aseptic Transfer

Aseptic transfer technique is important to prevent contamination of the culture being maintained as well as yourself. Manipulation of the tubes, plates and transfer tools requires patience and practice and is vital to success in the microbiology laboratory.

Proper use of a Bunsen burner: YouTube demo link title

Knowing how to adjust the Bunsen Burner flame for the most effective incineration is extremely useful. There are only two parts that you may need to adjust. Watch the YouTube video for a visual demonstration. Find the needle valve that controls the height of the flame on the bottom of the burner, check the integrity of the rubber tubing connecting the gas source to the gas inlet nozzle, and be sure to turn the gas on only when you are ready to light the burner and shut it off properly (at the gas source) whenever you are not using the flame.

Inoculation of a Liquid Culture: Broth to Broth, YouTube demo [[1]

1. Label the destination container for the culture.

2. Holding your loop like a pencil, insert the loop into the flame as illustrated in Figure A-1. The orientation of the loop wire in the flame is important for proper incineration. Keep the wire in the flame until it is red-hot. Then move the adjacent nonwire part of the loop lightly through the flame. The wire will now be sterile, and the nonwire part will have any dust burned off that might have fallen into the media during the transfer procedure. Remember that the loop is now hot and sterile. Allow the loop to cool for a few seconds in the air.

3. Pick up the stock broth tube of your organism with your other hand, while still holding the sterile loop. With the hand holding the loop, use your little finger against your palm to remove the cover from the culture tube as shown in Figure A-2. Do not put the cover down on your bench. If the tube was closed with a cotton or plastic plug, lightly pass the lip of the tube through the Bunsen burner to burn off any adhering dust. If the tube was closed with a plastic cap the lip of the tube should be sterile and this flaming is probably unnecessary. Now, insert the loop into the broth, and then remove it, carrying a loopful of culture. Flame the top of the culture tube, replace the tube cover, and return the tube to a rack.

4. Pick up the labeled destination tube or plate. Remove its cover, (flame if appropriate), insert the loop containing the culture into the broth, swirl gently and remove. Replace the cover and set the tube in the rack. Resterilize the loop before putting it down to avoid contamination of the bench with the culture. Be careful here. When the loop has liquid on it, you must insert the loop into the flame slowly to allow the liquid to evaporate rather than boil, which would splatter live bacterial cells all over the bench, your books and you.

5. Repeat steps 2, 3, and 5 for each liquid culture needed.

Slant to Broth, plate colony to broth: