2-4 Streak plates

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The streak plate method is a rapid and simple technique of mechanically diluting a relatively large concentration of microorganisms to a small, scattered population of cells. The goal is to obtain isolated colonies on a large part of the agar surface, so that desired species can then be brought into pure culture. Proper streaking of plates is an indispensable tool in microbiology. In most cases a closed inoculating loop is used for streaking plates. The wire loop should not be badly oxidized or pitted or it will fail to dilute the inoculum and will scratch the surface of the agar. Streak plates can be made from a broth culture, an agar slant or from an agar plate. It is sometimes convenient to suspend a bit of growth from a solid surface in sterile saline and use this as a source of inoculum. Resuspension of colonies or cultures grown on solid surfaces dilutes the culture and makes streak plating easier. A loopful of inoculum is transferred from the source and put on the agar surface. When using a large inoculum (a turbid culture or growth from a solid surface), a small spot is spread during the initial transfer. If the inoculum is from a lightly turbid suspension, the first phase of the streaking pattern is begun. Several basic patterns are illustrated in Figure 2-3. The three-phase streaking pattern is recommended for beginners because it is most likely to give satisfactory results with suspensions having a wide range of microbial density.

Figure 2-3 Streaking patterns

There are a number of different methods for mechanically diluting microbes on a streak plate. The most common method is spreading microbes across a plate as shown in the first four figures. As the concentration of microbes increases so do the number of phases. Irrespective of the number of phases, loop is flamed between each one. The fifth plate shows an alternative method, where the streaks are not continuous, but are a series of parallel lines. foobar

Choosing a streaking pattern is a matter of individual preference and depends upon the number of microorganisms in the sample. Figure 2-3 demonstrates the most common patterns, but they are not the only methods. The object of any streaking pattern is the continuous dilution of the inoculum to give many well isolated colonies. For multi-phase streaking it is crucial to flame the loop before starting the next phase. Note the slight overlap into the previous phase to pick up a small inoculum. To streak a plate...

  1. Flame the loop to sterilize it and let cool.
  2. Position the plate so that the spot of inoculum is nearest the hand not holding the loop (the opposite hand).
  3. Lift the plate lid with the opposite hand; just enough to get the loop inside and touch the loop to the inoculum spot. It is often helpful to treat the inoculating loop as if it were a pencil - steadying the loop by resting the heel of the hand against the lab bench.
  4. Move the loop back and forth across the spot and then gradually continue toward the center of the plate as you sweep back and forth. Use a very gentle and even pressure.
  5. When creating each phase, do not worry about keeping each pass across the plate separate from previous ones.
  6. When about 30% of the plate has been covered by the first streaking phase, remove the loop and flame sterilize it.
  7. Repeat the above procedure for the second phase, but this time pick up some inoculum by crossing into the first phase 2-3 times and then not passing into it again (Figure 2-3).
  8. Repeat as necessary for the third and fourth phases. After streaking the plate, flame sterilize the loop before setting it down.

Figure 2-4 demonstrates the technique of streak plates in a movie.

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