DSMZ 1512 . SCGYEM- MEDIUM

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Culture medium recipe

Vol.
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1 Medium for maintenance of infected amoeba cultures
Compound Amount Unit Conc. [g/L] Conc. [mM]
Solution A 800 ml - -
Solution B 50 ml - -
Solution C 50 ml - -
2 Combine Solutions A, B and C and add
Fetal bovine serum
(heat-inactivated, 56°C, 30 min)
100 ml - -
3 Maintenance conditions:
4 Infected amoeba cultures can be maintained e.g. in 25 cm2 flasks (e.g. Nunc flasks, VWR, 734-2081, Thermo-Scientific, 156340) at 20-30°C in an incubator (no special requirements concerning CO2 or light).
5 Maintenance/Passage of infected amoeba cultures:
6 Infected Acanthamoeba cultures don’t have to be transferred to fresh culture flasks as frequently as usually done for mammalian cell cultures. Amoeba cultures can be maintained in the same flask for several weeks or months. However, if the cultures reach a high cell density, medium should be exchanged. For that purpose amoebae are detached from the surface of the flask by knocking at the culture flask (no trypsin required, amoebae are only weakly adherent compared to most mammalian cells) and the medium (containing the majority of cells) is then completely removed. Fresh medium is added and the few remaining attached amoebae will continue to grow. Alternatively, cultures can also be passaged regularly (e.g. weekly) by transfer of a small aliquot of a densely grown culture (after cell detachment, as described above) to a fresh flask containing medium. If cultures contain few attached cells, but high amounts of cell debris, medium should be exchanged without prior cell detachment. Cultures usually recover fast. Depending on the host cell strain, the bacteria may cause host cell lysis. To maintain such cultures, bacteria have to be transferred by adding culture supernatant (approximately 0.5 ml) to fresh culture flasks containing uninfected host cells (approximately semi-confluent culture) each 7-10 days.
Equipment needed: Autoclave
Compound Amount Unit Conc. [g/L] Conc. [mM]
Casein 10.000 g 12.5 6.062
Na2HPO4 x 7 H2O 0.355 g 0.444 3.126
Distilled water 800.000 ml - -
1 Adjust pH to > 8.0, autoclave at 110°C
Equipment needed: Filter
Compound Amount Unit Conc. [g/L] Conc. [mM]
Glucose 2.5 g 50 277.531
KH2PO4 0.8 g 16 117.574
Distilled water 50.0 ml - -
1 Filter-sterilize
Equipment needed: Filter
Compound Amount Unit Conc. [g/L] Conc. [mM]
Yeast extract 5 g 100 -
Distilled water 50 ml - -
1 Filter-sterilize
Solution occurs in 2 other media (first defined in Medium 1500).
1 Bacteria released from host cells can be stored in
Compound Amount Unit Conc. [g/L] Conc. [mM]
Sucrose-Phosphate-Glutamate buffer 1000 ml - -
2 at -80°C for several months.
3 Infected cell cultures (supplemented with 10% DMSO) are best stored in liquid nitrogen.
Solution occurs in 2 other media (first defined in Medium 1500).
Compound Amount Unit Conc. [g/L] Conc. [mM]
Sucrose 75.00 g 75 219.106
KH2PO4 0.52 g 0.52 3.821
Na2HPO4 x 2 H2O 1.53 g 1.53 10.778
Glutamic acid 0.75 g 0.75 5.098
Distilled water 1000.00 ml - -

Metadata

Last modified: 19.12.21
Source: DSMZ
Taxonomic range:  Bacteria
Medium type: Complex medium
Final pH: n.d.
Equipment needed: Autoclave, Filter
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Associated strains

All strains for this medium: 2

Molecular composition

Compound Conc. [g/L] Conc. [mM]
Casein 10 4.85
Yeast extract 5 -
Glucose 2.5 13.877
KH2PO4 0.8 5.879
Na2HPO4 x 7 H2O 0.355 2.501
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Cultivation metadata from 2 strains BacDive

phyla 2 2 strains
temperature mesophilic 2 strains