Echinamoeba thermarum Baumgartner et al. (ATCC® PRA-13)

Organism: Echinamoeba thermarum Baumgartner et al.  /  Depositor: KO Stetter, M Baumgartner

Strain Designations OSB1
Application
Biotechnology
Biosafety Level 1
Isolation
hot spring
Wyoming, United States
Isolation date: June, 1995
Product Format frozen
Type Strain yes
Medium ATCC® Medium 2338: Echinamoeba broth
ATCC® Medium 2338: Echinamoeba broth
ATCC® Medium 2339: Echinamoeba agar
Growth Conditions
Temperature: 50.0°C
Growth conditions: Grown with Rhodobacter sp., strain OSrt, ATCC BAA-867
Cryopreservation
Cryoprotective Solution

DMSO                                                                                    1.5 ml

Fresh growth medium w/o bacteria                                 8.5 ml

1.     Mix the components in the order listed. When the medium is added to the DMSO the solution will warm up due to chemical heat.

2.    Harvest cells from a culture that is at or near peak density by filtration and centrifugation at 800 x g for 5 min.

3.  Adjust the concentration of cells at least 2 x 106/ml in freshmedium.

4.     Mix the cell preparation and the cryoprotective solution in equal portions.

5.     Dispense in 0.5 ml aliquots into 1.0 - 2.0 ml sterile plastic screw-capped cryules (special plastic vials for cryopreservation).

6.     Place vials in a controlled rate freezing unit. From room temperature cool at -1°C/min to -40°C. If freezing unit can compensate for the heat of fusion, maintain rate at -1 C/min through heat of fusion. At -40°C plunge ampules into liquid nitrogen. Alternatively, place the vials in a Nalgene 1°C freezing apparatus.  Place the apparatus at -80°C for 1.5 to 2 hours and then plunge ampules into liquid nitrogen.  (The cooling rate in this apparatus is approximately -1°C/min.)  

7.     Ampules are stored in either the vapor or liquid phase of a nitrogen refrigerator.

8.     To establish a culture from the frozen state place the vial in a 35°C water bath.  Immerse the vial to a level just above the surface of the frozen material. Do not agitate the vial.   Immediately after thawing, do not leave in water bath, aseptically remove the contents of the ampule and inoculate into a T-25 tissue culture flask containing 10 ml  bacterized ATCC medium 2338.  Screw the cap on tightly and incubate the culture at 50°C.  Alternatively, add the thawed contents of the ampule to the surface of a previously-bacterized Petri plate of ATCC medium 2338 agar.  Wrap the plate culture with parafilm and incubate upright at 50°C.

9.     Follow the protocol for maintenance of culture.

 

Name of Depositor KO Stetter, M Baumgartner
Special Collection National Park Service Special Collection (NPS)
Geographical Isolation Yellowstone National Park
Year of Origin June, 1995
References

Baumgartner M, et al. Cultivation and properties of Echinamoeba thermarum n. sp., an extremely thermophilic amoeba thriving in hot springs. Extremophiles 7: 267-274, 2003. PubMed: 12910386

outflow of Octopus Spring, White Creek, Yellowstone National Park, WY