Nutrient Agar Powder - Preparation & Equipment Use

1. Measure agar and distilled water into clean flask or beaker.

Recipe: Agar + Distilled Water = Yield  

Agar 23 g 11.5 g 9.2 g 4.6 g

+ Distilled Water 1000 ml 500 ml 400 ml 200 ml

= Yield 50 plates 25 plates 20 plates 10 plates

2. Flame sterilize a clean glass stir rod to stir the medium as it melts.
3. While wearing heat resistant hand protection, hold the flask or beaker over the flame. Swish or stir the mixture constantly while heating.
4. Boil the mixture for 1 minute. Remove from heat.
5. Place a sterile lab thermometer. in the mixture and monitor the temperature until it falls to approximately 45 - 50° C or if a lab thermometer is not available, cover and let stand a few minutes.
6. Pour enough melted agar into each sterile plastic petri dish to cover the bottom - about 1/8" to 1/4" deep. Replace the lid immediately.
7. Place agar plates on a counter top to cool and set. Agar medium will set like stiff gelatin at room temperature.
8. The agar medium is now ready for storage or use.

Preparing the Plates

1. If plates have been refrigerated, set them out and allow them to warm to room temperature.
2. Sterilize the loop
• To sterilize the loop, hold the handle with a pot holder and place the tiny looped wire in a flame until it turns bright red
• Allow the loop to cool for 3 - 5 seconds before touching the collection area.
• Resterilize the loop after each inoculation.
• Do not allow the loop to touch any surface other than the collection area and the agar.
3. Uncover each agar plate just long enough to inoculate the medium. hold the petri dish lid directly over the petri dish (or tilt the lid just enough to allow the loop inside) while inoculating the medium to help prevent contamination from air-borne particles. Do not allow the loop to touch the petri dish.
4. Do not Dip the loop in the agar, let it glide over the surface.
5. Make a pattern of inoculation lines (parallel lines, tic-tac-toe, zig-zag, initials, etc.) to help determine that what is growing is what you put there and not an air-borne contaminant.
6. Place the cover back on the plate immediately.

Incubation:

1. Turn the plates upside down and put them in a warm place. The ideal temperature for incubation is 32° C or 90° F. Bacterial growth should start to become visible in about 2 -3 days.

Science Fair Tips and Ideas for Kids:

1. Have someone take lots of pictures - some showing your face, some not showing your face.

Picture ideas:
• preparing materials for the experiment
• setting up to do the experiment
• doing the experiment, all the parts
• keeping a log of the experiment
• pictures of petri dishes with colonies
2. Keep a log of everything you do that relates to your project. Don't forget dates and times.
3. Label the bottom of each petri dish with a wax marker before inoculating them. Fewer mistakes are made when all the planning and preparation is done before starting the project.
4. Describe and number colonies. Measure them daily in mm. with a metric ruler. A clear ruler is ideal.
5. Use a hand lens or microscope to help in describing colonies.
6. List all your helpful resources; books, parents, teachers, web sites, and please don't forget us at Science Stuff.
7. Use color pictures and illustrations on your display board.
8. Be neat! A great project should have a great presentation! Measure and lightly mark where each item will go on your display board before you glue or tape it in place. Even with a great project, if your presentation is sloppy, you will lose points.