Biosafety classification is based on U.S. Public Health Service Guidelines, it is the responsibility of the customer to ensure that their facilities comply with biosafety regulations for their own country.
Frozen Cultures: -70°C for 1 week; liquid N2 vapor for long term storage
Freeze-dried Cultures: 2-8°C
Live Cultures: See Protocols section for handling information
Type Strain
no
Medium
ATCC® Medium 2454: Actinophrys medium
Growth Conditions
Temperature: 20°C to 25°C
Cryopreservation
Reagents
Cryoprotective Solution
DMSO 1.0 mL
Betaine 1.0 mL
Fresh growth medium 8.0 mL
Harvest and Preservation
Mix the components in the order listed. When the medium is added to the DMSO the solution will warm up due to chemical heat.
Harvest Actinophrys cells from a culture that has recently passed peak density by centrifugation at 300-400 x g for 5 min.
Adjust the concentration of cells to 1-2 x 104/mL in fresh medium.
Mix the cell preparation and the cryoprotective solution in equal portions by adding the cryoprotective solution to the cell suspension in 2 equal aliquots approximately 1 min apart.
Dispense in 0.5 mL aliquots into 1.0 - 2.0 mL sterile plastic screw-capped cryules (special plastic vials for cryopreservation).
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.)
Ampules are stored in either the vapor or liquid phase of a nitrogen refrigerator.
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 transfer to a petri plate or T-25 tissue culture flask optionally containing a bed of non-nutrient agar (ATCC medium 919) and 10 mL ATCC medium 2454
Aseptically transfer an appropriate volume of washed Chlorogonium to the petri plate or T-25 flask (see section on MAINTENANCE OF CULTURE). Incubate the culture at 20-25°C. Once the culture is established, follow the protocol for maintenance of culture.
Name of Depositor
T Suzaki
Special Collection
NSF - Protistology
Year of Origin
1994
References
Kinoshita E, et al. The ultrastructure of contractile tubules in the heliozoon Actinophrys sol and their possible involvement in rapid axopodial contraction. J. Eukaryot. Microbiol. 48: 519-526, 2001. PubMed: 11596916
Sakaguchi M, Suzaki T. Monoxenic culture of the heliozoon Actinophrys sol. Eur. J. Protistol. 35: 411-415, 1999.
