Aseptic Technique

Techniques to keep a sterile environment in a micro lab.

What is Aseptic Technique?

Aseptic technique refers to a procedure that is performed under sterile conditions.


To minimize the risk of contamination, a method known as aseptic technique is practiced. Aseptic technique is a procedure designed to keep unwanted microorganisms from contaminating sterile materials or pure cultures of microorganisms. Not only can unwanted species contaminate a pure culture and yield incorrect results but they also may be pathogenic and facilitate disease growth. Therefore the prudent practice of aseptic technique is extremely important when working in the microbiology laboratory.

Plate Streaking

Most source materials contain a large number of microorganisms. Pond water can be expected to contain 105 to 106 bacteria/ml , soil to contain more than 108 bacteria per ml, and a barely visibly turbid culture of Escherichia coli contains at least 107 bacteria per ml. This means that a loop of these materials spread over the surface of an agar plate would yield 103 to 105 colonies, producing a confluent lawn of growth on the plate. Thus, simply smearing a loop of source material over a plate will not yield isolated colonies. Additional steps are needed.

Steps for a Three-Phase Streak (how to isolate an organism)


  • To streak a sample for isolation, draw a "T" on the back of the plate, dividing it into three sections.
  • Flame the loop, allow it to cool a few seconds, and then pick up a loop of sample. Turn the plate so that the top of the "T" is near the palm of your non-dominant hand.
  • Crack the lid of the plate by lifting it with your thumb and index finger.
  • Beginning at the edge of the plate above the top of the "T" and move the loop back and forth across the agar surface, pulling it towards the top of the "T" as you do so.

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  • Remove the loop, flame-sterilize, and cool it. Do NOT insert the loop back into the sample.
  • Turn the plate 90 degrees counter-clockwise.
  • Repeat the streaking procedure, beginning at the edge of the plate. This time, cross the previous material three or four times to pick up cells from the part you first streaked.
  • Continue streaking until the loop reaches the bottom of the top of the "T."
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  • Remove the loop, flame-sterilize, and cool it. Do NOT insert the loop back into the sample.
  • Turn the plate 90 degrees counter-clockwise.
  • Repeat the streaking procedure, beginning at the edge of the plate. Again, cross the previous material three or four times to pick up cells from the part you streaked second.
  • Continue streaking until the loop reaches the bottom of the top of the "T."
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Micro eGuide - Streak Plate Technique

Broth Transfer Technique

  • Loosen the test tube cap, if necessary, but do not remove it.
  • Hold the loop in your dominant hand and flame-sterilize it. Allow the loop to cool a few seconds.
  • Pick up the test tube in your non-dominant hand. Remove the cap from the test tube. Do not set the cap down.
  • Insert the loop into the tube and pick up a loopful of culture.
  • Remove the loop and carefully replace the cap on the tube.
  • Return the tube to the rack.
  • Pick up the tube you wish to inoculate.
  • Remove the lid.
  • Slide the loop into the tube. (You do not need to rattle the loop to shake off the bacteria.)
  • Remove the loop and carefully replace the cap on the tube.
  • Return the tube to the rack.
  • Flame the loop and set it down.
Micro eGuide - Tube Transfers

Pipette Transfer


  • Pick up a pipette with your non-dominant hand. Use your dominant hand to place a pipette aid on the pipette.
  • Slide the pipette from the con,tainer lifting it to avoid dragging the bottom of the pipette across the contaminated surfaces of the other pipettes.
  • Use the little finger of the hand holding the pipette to remove the lid from the container containing your sample.
  • Touch the tip of the pipette to the side of the container.
  • Measure sample into the container.
  • Place the pipette into a discard container and remove the pipette aid.
Micro eGuide - Aseptic Techniques: Pipette Transfers

Flaming the Loop

  • If the loop has been dipped in culture since it was last flamed, hold it above the Bunsen Burner flame to slowly dry the liquid it contains. This will reduce spattering and the production of aerosols when the loop is sterilized.
  • Hold the loop with the tip in the hottest part of the flame (just above the bright blue cone). Holding the loop at a fairly steep angle will allow you to sterilize more of its length at one time.
  • Allow the loop to cool a few seconds before placing it in any agar or broth.