Table 4.

Medium for in vitro growth of G. lambliaa

ComponentQuantity/literFinal concn
K2HPO4 · H2O1 g4.4 mM
KH2PO4600 m4.4 mM
Trypticase (casein peptone)20 g
Yeast extract10 g
d-Glucose10 g56 mM
NaCl2 g34 mM
Cysteine-HCl2 g16.5 mM
Ascorbic acid200 mg1.1 mM
Ferric NH4-citrate (Sigma F-5879)22.8 mg
Bovine bile (Sigma B-8381)40 ml of 6.5% solution
NaOHBring pH to 7.0–7.2 (1.65 ml of 10 M solution
Serum (bovine or fetal calf)100 ml10%
  • a Formula of TYI-S-33 (159), the most commonly used medium for growth of G. lambliatrophozoites. The ferric-NH4 citrate and bovine bile are generally kept as solutions at 4°C. All components except the serum are dissolved in water, brought to 200 ml, sterilized on a 0.22- or 0.45-μm-pore-size filter, and added to 700 ml of sterilized water. Heat-inactivated (56°C for 20 min) serum is then added. During in vitro growth, the organisms are grown in sealed glass containers nearly filled with medium. When this is impossible, such as during cloning by limiting dilution in 96-well plates (243), a sealed bag containing an anaerobic generator is used. At 4°C, the shelf life is limited to 5 to 7 days, primarily because of the degradation of the cysteine.