Diplomonads:Giardia Medium

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If you have a different media protocol, add it as another header here.

Phosphate-buffered Giardia Growth Medium

5X Basic Medium (500ml)

in about 400ml milliQ water add:

• 25 g D-glucose
• 5g NaCl
• 25 g yeast extract
• 50 g casein peptone digest (SIGMA P6838, Lot#27H1113)

1. Stir on stirplate for about 30 minutes.
2. Bring to 500ml volume with more milliQ water.
3. Prefilter with a 0.22µm filter. This keeps >4 months at 4°C.

Complete Medium (500ml)

Thaw ABS (adult bovine serum), FAC (ferric ammonium citrate) and PSF (PenStrepFungizone) in 37C water bath.

Then in a 500ml beaker, add:

• 350 ml milliQ water
• 1 g cysteine (SIGMA C-7880)
• 0.05 g ascorbic acid

1. Stir to mix, and pH to 7.0-7.2 with 10M NaOH Transfer to 500ml purple cap media bottle.
2. In laminar flow hood, add the following to the cysteine solution from above:

• 10 ml 50X Phosphate Buffer Solution (50X= (15g KH₂PO₄/32.5g K₂HPO₄x3H₂O in 500ml)
• 100 ml semi-sterile basic medium (see above) ***make sure to warm semi-sterile basic medium to 37 degrees C for 30 min, or else it will clog the filter***
• 4 ml sterile 6.5% bovine bile (stored at 4°C)
• 1.5 ml sterile FAC (ferric ammonium citrate) (2.28mg/ml stock)
• 5 ml pen/strep+fungizone (Gibco)
• 50 ml Adult Bovine Serum (Invitrogen, lot #)

1. Shake to mix in bottle, then filter with 0.22µm filter. Medium should be clear and amber-colored. The medium only lasts about 7-10 days, so it is best make it fresh every week. Cysteine precipitates out of solution easily over time (looks like crystals in microscope or flakes in bottle). If this happens, change medium immediately.
2. Store at 4C.