Tons of microbiology experiments rely on the proper growth of bacteria on media plates or suspension cultures. To have optimized growth conditions, two things are primarily important: Aseptic lab conditions and knowledge of bacterial inoculation. As for the third important thing to have successful bacterial growth, good quality media plates are required and Advancells can help you with that!
Here we discuss the most common method of bacterial inoculation on agar plates: Streaking. But first, let us see how to prepare a sterile workplace:
- Know the laboratory rules and follow safety guidelines in accordance with institutional Environmental Health and Safety departments. Have proper knowledge of waste and biohazard disposal.
- Sterilize instruments and work prior to using them.
- Clean clutter in the work area or laboratory bench.
- Clean work area with disinfectant.
- Set up Bunsen burner and work carefully within the sterile field area.
- Arrange all the supplies for the procedure on the laboratory bench and make sure to label the materials. Organizing the work area maximizes work efficiency by minimizing the exposure time of materials to airborne contaminants.
- Once settled comfortably with the properly organized arrangement in the working area, slightly loosen the caps of tubes, flasks, and bottles to open them easily with one hand during subsequent handling.
- Wash hands with antiseptic soap and warm water, and let them dry before handling microorganisms.
Streak Plate Procedure
The streaking procedure is designed to isolate pure cultures of bacteria from mixed populations by simple separation using the quadrant method. Single colonies (visible) comprise millions of bacteria cells in a cluster on agar plates. Theoretically, cells in a colony are derived from a single bacterium and therefore can be referred to as a clonal cluster of genetically identical cells.
With the streak-plate procedure, a cell suspension droplet is spread over the surface of the agar-based media plate so that fewer bacterial cells are deposited at widely separated points. This helps the information of individual separated colonies upon incubation. Streak-plating can be done generally using metal loops or disposable plastic loops. A metal loop can be re-used while many researchers prefer pre-sterilized disposable wooden sticks or plastic loops for streak-plating, as an inexpensive alternative.
Some important things to remember:
- The label around the edge of the plate bottom with name initials, date, type of growth medium, and type of bacteria. Avoid doing that on the lid as it can be misplaced.
- Plates must be completely dry without condensation on the lid. They should also be pre-warmed to room temperature before streak-plating. If the media plates are stored at 4 °C, ensure that they are removed several hours before to attain room temperature.
- The bacteria sample used for streak-plating could be either a suspension or an existing colony from another agar plate. It is recommended that only one type of bacterial sample is used for individual plate inoculation, as a beginner. For a professional microbiologist, it is okay to use multiple samples for a single plate.
Ways to inoculate from suspension or another colony:
- If the inoculum is a cell suspension, vortex it before removing an aliquot for plating. Flame the loop and the bottle or tube rim, before and after inoculating. Do not touch the loop with tube or bottle outer surface. After flaming the loop, wait for 2-5 seconds before touching the fresh agar surface (this ensures that the loop is not too hot) and then dip the loop in suspension. Following that, place the loop near the end of one quadrant and streak all the way down in a zig-zag manner.
- If the inoculum is a colony from another plate, we do not have to transfer the whole colony. The process is similar to the previous section but after you touch the sterile loop on the agar surface to ensure that the loop is not too hot, slightly scrape on the previous colony and streak it on the quadrant.
- Always ensure that the streak of one quadrant does not overlap with the streak of another quadrant.
- Incubate the plate upside down so condensation that accumulates on the lid does not drip on the colonies.
There are other methods of inoculating like spread-plate and pour-plate but every method of plating has its own purpose in research. Contact firstname.lastname@example.org if you are looking for ready-to-use agar plates for your microbiology research.