How to make prepared agar media



Pre-experiment: Keep sterile Petri dishes closed until ready to pour agar into them. Air-borne contaminants can easily invade an open Petri dish.

Although pre-poured agar plates are available, one can make agar plates from tablet, powdered, or bottled agar by following a few simple instructions. Agar kits usually come with detailed instructions on how to prepare plates, and below are sample procedures for reference. When in doubt, be sure to clearly read the instructions and ask for help if needed (either consult a teacher or call the technical help line of the agar kit supplier).

Preparing Tablet or Powdered Agar:
For example : The formulation for LB (Luria Bertani) agar is: 9.1 g/L tryptone, 4.6 g/L yeast extract, 4.6 g/L NaCl, and 13.7 g/L agar. If using tablets, dissolve 10 tablets per 500 ml of water. For agar powders, dissolve by microwaving, 6.9 g of agar in 500 ml of water. 500 ml of agar will pour ~ 25 large Petri dishes (100 mm diameter) or 50 small Petri dishes (60 mm diameter).

Preparing Bottled Agar:
Loosen the bottle cap, but do not remove the cap while heating.
Warm the agar bottle in a hot water bath or in the microwave until it becomes liquid.
After opening the cap, pass the neck of the agar bottle through a flame to sterilize it. Do not lose the cap!
While pouring the agar, open the Petri dish lid as little as possible, hold it at an angle, and make sure the lid is kept directly over the Petri dish.
Pour enough melted agar into each sterile plastic Petri dish to cover 1/8" of the bottom. Cover the lid of the Petri dish immediately.
Place agar plates on a counter top to cool and set. Agar medium will set like stiff gelatin at room temperature.
Pass the neck of the agar bottle through flame again before applying the cap.
Preparing Pre-Poured Plates: If plates have been refrigerated, set them out and allow them to warm to room temperature.

Storage: Stack agar plates upside down in the refrigerator. Do Not Freeze! The purpose of placing the plates upside down is to prevent condensation from dripping down onto the agar surface which could then facilitate movement of organisms between colonies.
Please see http://www.umsl.edu/~microbes/pdf/tipsforplates.pdf for additional tips.

Additional Safety Considerations
When stirring the broth solution, one should take special note in beginning the stir scale at a low setting and adding more speed from there.
When heating the broth, make sure to cover the flask in such a manner that will not lend itself to boiling over, but to avoid spillage.
When pouring the broth, make sure to fill the Petri dish without burning oneself. In addition it is important in this process to make sure that the Petri dish is covered immediately to allow the substance to cool proportionately.
Once the Petri dishes have been exposed or inoculated, students should not re-open them.

0 Response to "How to make prepared agar media"

Post a Comment