Acanthamoeba royreba Willaert et al. (ATCC® 30884)

Organism: Acanthamoeba royreba Willaert et al.  /  Depositor: E Willaert

Permits and Restrictions

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Strain Designations Oak Ridge [AC-023]
Application
characterization of Acanthamoeba polyphaga
Biosafety Level 1
Isolation
BeWo human choriocarcinoma cells, Oak Ridge, TN, 1975
Product Format freeze-dried
Type Strain yes
Comments
Taxonomy based on isoenzyme profiles and rDNA PCR-RFLP patterns
species description
characterization of Acanthamoeba polyphaga
review
phylogeny
Medium Medium 712: PYG w/ Additives
Growth Conditions
Temperature: 25.0°C
Duration: axenic
Protocol: ATCCNO: 30010 SPEC: This strain is distributed as a freeze-dried preparation. See the general procedures for opening a freeze-dried vial. Aseptically add 0.5 ml of ice cold medium containing 12% (w/v) sucrose to the freeze-dried inner shell vial. Once the culture is completely rehydrated, aseptically add 1 ml of ATCC medium 712 and distribute to a 16 X 125 mm plastic screw-capped test tube or a T-25 tissue culture flask containing 5.0 ml of the same medium. Incubate the test tube culture horizontally with the cap on tight. Trophozoites should be evident in 1-5 days.
Subcultivation
Protocol: ATCCNO: 30010 SPEC: This strain is distributed as a freeze-dried preparation. See the general procedures for opening a freeze-dried vial. Aseptically add 0.5 ml of ice cold medium containing 12% (w/v) sucrose to the freeze-dried inner shell vial. Once the culture is completely rehydrated, aseptically add 1 ml of ATCC medium 712 and distribute to a 16 X 125 mm plastic screw-capped test tube or a T-25 tissue culture flask containing 5.0 ml of the same medium. Incubate the test tube culture horizontally with the cap on tight. Trophozoites should be evident in 1-5 days.
Cryopreservation

1.   To achieve the best results set up cultures with several different inocula (e.g. 0.25 ml, 0.5 ml, 1.0 ml).  Harvest cultures and pool when the culture that received the lowest inoculum is at or near peak density.

2.  If the cell concentration exceeds the required level do not centrifuge, but adjust the concentration to between 2 x 106 and 2 x 107cysts/ml with fresh medium.  If the concentration is too low, centrifuge at 600 x g for 5 min and resuspend the pellet in the volume of fresh medium required to yield the desired concentration.

3.  While cells are centrifuging prepare a 15% (v/v) solution of sterile DMSO as follows:  Add the required volume of DMSO to a glass screw-capped test tube and place it in an ice bath.  Allow the DMSO to solidify.  Add the required volume of refrigerated medium.  Dissolve the DMSO by inverting the tube several times. 

      *NOTE: If the DMSO solution is not prepared on ice, an exothermic reaction will occur that may precipitate certain components of the medium.

4.  Mix the cell preparation and the DMSO in equal portions. Thus, the final concentration will be between 106 and 107 cells/ml and 7.5% (v/v) DMSO. The time from the mixing of the cell preparation and DMSO stock solution before the freezing process is begun should be no less than 15 min and no longer than 30 min.

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 the vials in a controlled rate freezing unit.  From room temperature cool at -1°C/min to -40°C.  If the freezing unit can compensate for the heat of fusion, maintain rate at        -1°C/min through the heat of fusion.  At -40°C plunge 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. The frozen preparations are stored in either the vapor or liquid phase of a nitrogen freezer.

8.   To establish a culture from the frozen state place an ampule in a water bath set at 35°C (2-3 min). Immerse the vial just sufficient to cover the frozen material. Do not agitate the vial.

9.   Immediately after thawing, aseptically remove the contents of the ampule and inoculate into 5 ml of fresh ATCC medium 712 in a T-25 tissue culture flask or plastic 16 x 125 mm screw-capped test tube.  Incubate at 25°C.

Name of Depositor E Willaert
Chain of Custody
ATCC <<--E Willaert<<--R.L. Tyndall
Year of Origin 1975
References

Willaert E, et al. Acanthamoeba royreba sp. n. from a human tumor cell culture. J. Protozool. 25: 1-14, 1978. PubMed: 566323

Tyndall RL, et al. Pathogenic and enzymatic characteristics of Acanthamoeba isolated from cultured tumor cells. Protistologica 15: 17-22, 1979.

Hall J, Voelz H. Bacterial endosymbionts of Acanthamoeba sp.. J. Parasitol. 71: 89-95, 1985. PubMed: 3981353

Moura H, et al. Acanthamoeba healyi n. sp. and the isoenzyme and immunoblot profiles of Acanthamoeba spp., groups 1 and 3. J. Protozool. 39: 573-583, 1992. PubMed: 1522539

John DTOpportunistically pathogenic free-living amebaeIn: John DTParasitic protozoa2nd ed.3San DiegoAcademic Presspp. 143-246, 1993

Daggett PM, et al. Distribution and possible interrelationships of pathogenic and nonpathogenic Acanthamoeba from aquatic environments. Microb. Ecol. 8: 371-386, 1982.

Daggett PM, et al. A molecular approach to the phylogeny of Acanthamoeba. Biosystems 18: 399-405, 1985. PubMed: 4084681

Kim YH, et al. Close relatedness of Acanthamoeba pustulosa with Acanthamoeba palestinensis based on isoenzyme profiles and rDNA PCR-RFLP patterns. Korean J. Parasitol. 34: 259-266, 1996. PubMed: 9017912

type strain

Alves JMP, et al. Random amplified polymorphic DNA probes as a tool for the characterization of Brazilian keratitis isolates of the genus Acanthamoeba. Braz. J. Med. Biol. Res. 33: 19-26, 2000.

Marciano-Cabral F, Cabral G. Acanthamoeba spp. as agents of disease in humans. Clin. Microbiol. Rev. 16: 273-307, 2003. PubMed: 12692099

Flint JA, et al. Genetic analysis of forty isolates of Acanthamoeba Group III by multilocus isoenzyme electrophoresis. Acta Protozool. 42: 317-324, 2003.

Cross References

Nucleotide (GenBank) : U07417 nuclear SSU rRNA gene

Notice: Necessary PermitsPermits

These permits may be required for shipping this product:

  • Customers located in the state of Hawaii will need to contact the Hawaii Department of Agriculture to determine if an Import Permit is required. A copy of the permit or documentation that a permit is not required must be sent to ATCC in advance of shipment.
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