Tan Chaos

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.