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Preparing Agar Plates

Preparing Agar Plates. As demonstrated by Andrew Corcoran. Use an appropriate flask. Use a flask that is twice the volume of the media you will make Typically a 1 L flask to prepare 0.5 L batches. Measure out the water . Add a spin bar and spin. Weigh out the appropriate amount of media.

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Preparing Agar Plates

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  1. Preparing Agar Plates As demonstrated by Andrew Corcoran

  2. Use an appropriate flask • Use a flask that is twice the volume of the media you will make • Typically a 1 L flask to prepare 0.5 L batches

  3. Measure out the water

  4. Add a spin bar and spin

  5. Weigh out the appropriate amount of media

  6. Add the powder • Some powders are very fine

  7. Make sure all the powder gets hydrated Wad of dry media

  8. Heat until there is a hint of boiling • Avoid boiling over! • Agar will partially dissolve and get a little clearer • Wear a “Clavie”

  9. Remove the spin bar with the wand Wand Spin Bar

  10. Apply an aluminum foil lid

  11. Use autoclave tape • Turns color after heating • Tear off tape and apply to counter (easier to write on it) • Include initials and media type

  12. Apply to flask

  13. Always place media into autoclave tray • Secondary containment • Spills in the autoclave clog vents

  14. Operating autoclave Jacket Pressure - There should always be pressure here Chamber pressure – Should read zero • First rule – always make sure there is no pressure in the autoclave!

  15. Make sure previous cycle is finished Chamber pressure – Should read zero Autoclave status

  16. Open the door Pins • Lefty loosey • Pins will retract • Crack 1 inch. Allow excess steam to vent • Open slowly • Remove contents carefully (contents may be superheated)

  17. Place tray in center of rack Shelves • Slide out metal shelf (avoid reaching into autoclave)

  18. Close the door • Righty tighty • Make sure pins engage • Door closed light will be lit • Turn until hand tight

  19. Press flask icon • Fast exhaust – Scissors • Fast exhaust with drying – Gown • Slow exhaust - Flask

  20. The control panel Fast exhaust Fast exhaust/dry Slow exhaust Service – don’t touch Cycles are pre-programmed. There is no need to change settings! Start button

  21. Press start

  22. Steam pressure will rise

  23. After autoclaving place flask in 55 oC waterbath Tape has darkened

  24. Pouring the plates • Start by disinfecting the work area

  25. Stack plates 4 high (right side up)

  26. Label plates on the bottom • Media type • Date • Label the bag they will go in

  27. Wipe away water from flask

  28. Remove lid and begin pouring plates from the bottom of the stack

  29. After the first plate is filled cover and begin filling the second (and so on)

  30. Avoid moving or jostling plates Condensation will form on lid of plate Agar on the lid - faux pas!

  31. When done, put hot water in flask

  32. Pour into special trash can

  33. Stack the plates • This prevents condensation on most plates • Also provides more work room

  34. Any questions?

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