Tan Chaos

Tan Chaos kdorfman Thu, 02/16/2012 - 14:27

See protocol on Chaos main page

Maintenance

  • Use culture-treated dishes
  • keep in Chaos saline at 22-24C in the dark
  • Every 2-3 days
    • count
    • change medium
    • feed
  • Split every 2-3 weeks, or when the debris becomes detrimental

Feeding

  • 7-day old Paramecium multimicronucleatum cultures at late log phase or early stationary phase (Fok and Allen, 1979).
  • Remove growth medium:
    • Filter cultures through a 3-piece glass filter funnel (Whatman)

http://www.whatman.com/GlassMicrofiberAccessories.aspx 1950-007 Filter Funnel, 3-piece, 70 mm, 115 ml reservoir 1/pk Fisher 09-877-3 $223.58

lined with a layer of GF/A glass microfibre filter (Whatman) 7.0cm dia. 1820-070
Fisher 09-874-16A Pack of 100 for $48.50

  • Wash and concentrate Paramecia in Chaos saline

    • concentrate by allowing the medium to be filtered through
    • collect concentrated Paramecium suspension in a beaker

  • Estimate Paramecium concentration

    • 10 10uL drops
    • count # Paramecium in each
    • average x 100 = cells/mL

  • Feed 36 ± 3 Paramecia per amoeba.

Changing the medium

  • Tilt the dish, let amoebae settle to the bottom
  • Suck out the old medium
  • Carefully squirt fresh medium onto the dish surface
  • Repeat 2 or 3 times
  • Check the culture
    • Fast growing and healthy cultures have little cell debris (dead amoebae, cell fragments) and usually few or no lingering Paramecium left from the previous feeding.
    • slow-growing or sick cultures have more cell debris and a large number of Paramecium left from the previous feeding.