nahrein.yaghoubpour wrote:Thanks Lisa for replying. I thought it might be the amount of agar I used or the agar being old.
Agar agar has a fairly infinite shelf-life; just so long as it's kept dry.
To make a litre, disolve 15-20g agar agar, and nutrients, in about 850mL of cold distilled water. (Eg: 5g peptone, 3g beef extract or substitutes such as vegemite.)
Mix thoroughly until you get a cloudy suspension, with no powder settling on the bottom or floating on the top.
Bring to near boiling while stirring for about five minutes. Don't allow to boil; add cold distilled water to keep the brew from bubbling. Continue until the solution clarrifies into a
clean yellow solution. It won't go fully clear, but the phase transition is noticable. This is your visual test. If it's still cloudy it won't set.
Optionally at this point, pipette a
test dash into the dimple of a cold dropping tile and pop into the fridge for a moment to see if it will set properly. Just continue heating, and maybe disolve in a few more grams of agar agar, until satified.
Let the solution start to cool, then dilute to volume with cool distilled water.
Pour into conical flasks with a cotton plug, capped by aluminium foil, and autoclave for 15 minutes at 140 kPa. These can be stored in the fridge until needed (and remelted in the autoclave again for a few minutes), or used immediately.
However, the autoclaved solution should be allowed to cool to below 50 C before pouring onto plates, to reduce condensation. Additionally, this means the conical flasks should be cool enough to handle by hand rather than needing clumby heat proof gloves. (But do test carefully.)
