Viable Counts: Plate Streaking

The invention of solid media for the growth of bacteria made it easier to count bacteria by plating samples. Diluted samples can be spread on the surface of solidified agar plates or mixed into melted agar. When the bacteria reproduce, they produce distinct colonies. It is assumed that each colony represents one viable cell from the inoculant.

Spread Plates

  • Obtain a Bunsen burner, a container of alcohol, and a bent glass rod (also known as a dally or hockey stick).
  • Label the bottoms of Petri plates containing medium with the sample, date, your name, and your class.
  • Place the dally into the container of alcohol and light the Bunsen burner.
  • Aseptically draw sample into the pipette2 using a pipette aid.
  • Crack the lid of the Petri plate using your non-dominant hand.
  • Insert the pipette under the Petri plate lid so that the tip of the pipette touches the agar surface. Holding the pipette fairly horizontally will make the flow of liquid easier to control.
  • Pipette sample (usually 0.1 ml) onto the agar surface.
  • Place the pipette into a discard container and remove the pipette aid.
  • Immediately pass the dally through the Bunsen burn flame to light the alcohol that clings to it. Do not hold the dally in the flame.
  • Allow the alcohol to burn off.
  • Cracking the lid of the Petri plate with your non-dominant hand, use the dally to spread the sample evenly over the surface of the entire plate.
  • Rotate the plate 90 degrees, and continue spreading the sample.
  • Place the dally back into the alcohol.
  • Incubate the plate agar side up as directed.

Pour Plates

  • Melt agar medium by boiling, steaming, or autoclaving, and cool it in a 50o C water bath.
  • Label the bottoms of empty Petri plates with the sample, date, your name, and your class.
  • Aseptically draw sample into the pipette1 using a pipette aid.
  • Crack the lid of the Petri plate using your non-dominant hand.
  • Insert the pipette under the Petri plate lid so that the tip of the pipette touches the bottom of the plate. Holding the pipette fairly horizontally will make the flow of liquid easier to control.
  • Pipette sample (usually 1.0 or 0.1 ml) onto the bottom of the plate.
  • Place the pipette into a discard container and remove the pipette aid.
  • Remove the container of agar from the water bath and wipe the water from the outside of the container with a paper towel.
  • If your bottle is made of glass, briefly pass the mouth of the container through the flame so that agar will not be contaminated when you pour it out. Skip this step if your bottle is made of plastic or there is plastic near the lid of your bottle.
  • Pour agar into the plate, covering 1/2 to 2/3 of the bottom of the plate.
  • Immediately mix the sample into the agar by gently swirling the plate on the countertop in a figure eight pattern.
  • Allow the plate to cool until the agar has solidified.
  • Incubate the plate agar side up as directed.

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