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BISC209: Aseptic Transfer: Difference between revisions
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'''Aseptic transfer''' <br> | '''Aseptic transfer''' <br> | ||
'''Broth to Broth'''<BR> | '''Broth to Broth or Broth to Plate Transfer'''<BR> | ||
YouTube demo [http://www.youtube.com/watch?v=0odxJy0nR9s&feature=PlayList&p=4DA5158C3E7E14AC&playnext=1&playnext_from=PL&index=20] | YouTube demo [http://www.youtube.com/watch?v=0odxJy0nR9s&feature=PlayList&p=4DA5158C3E7E14AC&playnext=1&playnext_from=PL&index=20] | ||
1. Label the destination container for the culture. | 1. Label the destination container for the culture (uninoculated sterile broth in a tube or solid medium in a plate). | ||
2. Holding your loop like a pencil, insert the loop into the flame as illustrated in Figure 1. The orientation of the loop wire in the flame should be at ~ a 30 degree angle 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. Allow the loop to cool for a few seconds in the air before touching it to your culture or medium. <br> | 2. Holding your loop like a pencil, insert the loop into the flame as illustrated in Figure 1. The orientation of the loop wire in the flame should be at ~ a 30 degree angle 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. Allow the loop to cool for a few seconds in the air before touching it to your culture or medium. <br> | ||
[[Image:Asep1_0.75.jpg]] <br> | [[Image:Asep1_0.75.jpg]] <br> | ||
Figure 1: Proper flaming of a loop. Note how the loop handle is held by only the thumb and first two fingers and the loop is inserted into the hottest part of the flame. | ''Figure 1: Proper flaming of a loop. Note how the loop handle is held by only the thumb and first two fingers and the loop is inserted into the hottest part of the flame.'' | ||
3. Pick up the donor broth culture tube 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 or plug from the culture tube as shown in Figure 2. Do not put the cover or plug down on your bench. If the tube is glass, lightly pass the lip of the tube through the Bunsen burner to burn off any adhering dust and to create a temperature differential that temporarilty prevents dust from falling into your tube. Now, insert the loop into the broth, and then remove it, carrying a loopful of culture. | 3. Pick up the donor broth culture tube 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 or plug from the culture tube as shown in Figure 2. Do not put the cover or plug down on your bench. If the tube is glass, lightly pass the lip of the tube through the Bunsen burner to burn off any adhering dust and to create a temperature differential that temporarilty prevents dust from falling into your tube. Now, insert the loop into the broth without touching the sides of the tube, and then remove it, carrying a loopful of culture. Pass the top of the culture tube through the flame, replace the tube cover or plug, and return the tube to a rack.<br> | ||
[[Image:Asep2.jpg]]<br> | [[Image:Asep2.jpg]]<br> | ||
Figure 2: Transferring a culture. (a) Removal of a tube cap while manipulating a loop; (b) Obtaining inoculum from a broth tube while maintaining sterility of the cap (note cap in hand).<br> | ''Figure 2: Transferring a culture. (a) Removal of a tube cap while manipulating a loop; (b) Obtaining inoculum from a broth tube while maintaining sterility of the cap (note cap in hand).''<br> | ||
4. Pick up the labeled destination tube or plate. Remove its cover, (flame | 4. Pick up the labeled sterile destination tube or plate. Remove its cover, (if it's a glass tube, pass the lip of the tube through the flame),<BR> | ||
5. Insert the loop containing the culture into the broth, swirl gently and remove. | 5. Insert the loop containing the culture into the destination tube of sterile broth, swirl gently and remove. If the destination is a plate of solid medium, follow the directions for streaking for isolation or for a spread plate found in the protocols section of the wiki. <br> | ||
6. Replace the cover and set the tube in the rack. <br> | 6. Replace the cover and set the newly inoculated broth tube in the rack. <br> | ||
7. Resterilize the loop before putting it down | 7. Resterilize the loop before putting it down by inserting the loop into the flame ''very slowly''. Doing this slowly allows any liquid remaining on the loop to evaporate rather than boil and avoid splattering live bacterial cells all over the bench and you.<br> | ||
'''Inoculation of a Slant'''<br> | '''Inoculation of a Slant'''<br> | ||
