Acanthamoeba polyphaga (Puschkarew) Page (ATCC® 30461)

Strain Designations: eye  /  Depositor: GS Visvesvara  /  Biosafety Level: 2

Permits and Restrictions

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Strain Designations eye
Host for pathogenic Legionella species
Characterization of Acanthamoeba polyphaga
Characterization of proteolytic secretions
Biosafety Level 2

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.

Isolation Human corneal scrapings, Houston, TX, 1973
Product Format frozen
Storage Conditions Frozen Cultures:
-70°C for 1 week; liquid N2 vapor for long term storage

Freeze-dried Cultures:

Live Cultures:
See Protocols section for handling information
Type Strain no
Antibiotic Resistance
resistant to complement lysis
In vitro effectiveness of povidone-iodine
Host for pathogenic Legionella species
Resistant to complement lysis
Characterization of Acanthamoeba polyphaga
Specific antigens revealed by protein immunoblotting
Mitochondrial DNA fingerprinting
Effects of azole compounds
Use of plastic ampoules for freeze preservation
Characterization of proteolytic secretions
Inhibition by Pseudomonas aeruginosa
Quantitative bacterial plaque assay for enumeration
Differentiation of Naegleria fowleri from Acanthamoeba using monocolonal antibody
Internalisation of bacterial substrates as factors in growth rate
Proteinase activities
Medium ATCC® Medium 712: PYG w/ Additives
Growth Conditions
Temperature: 25°C
Culture System: Axenic
Cryopreservation Harvest and Preservation
  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 mL to 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 to 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 GS Visvesvara
Chain of Custody
ATCC <-- GS Visvesvara <-- D.B. Jones/N.M. Robinson
Year of Origin 1973

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Kong HH, et al. Interstrain polymorphisms of isoenzyme profiles and mitochondrial DNA fingerprints among seven strains assigned to Acanthamoeba polyphaga. Korean J. Parasitol. 33: 331-340, 1995. PubMed: 8591011

Kong HH, et al. Mitochondrial DNA restriction fragment length polymorphism (RFLP) and 18S small-subunit ribosomal DNA PCR-RFLP analyses of Acanthamoeba isolated from contact lens storage cases of residents in southwestern Korea. J. Clin. Microbiol. 40: 1199-1206, 2002. PubMed: 11923331

Lai S, et al. Non-radioactive DNA probe and polymerase chain reaction procedures for the specific detection of Acanthamoeba. Mol. Cell. Probes 8: 81-89, 1994. PubMed: 8028612

Marciano-Cabral F, Toney DM. The interacion of Acanthamoeba spp. with activated macrophages and with macrophage cell lines. J. Eukaryot. Microbiol. 45: 452-458, 1998. PubMed: 9703682

Mitro K, et al. Partial characterization of the proteolytic secretions of Acanthamoeba polyphaga. Exp. Parasitol. 78: 377-385, 1994. PubMed: 8206136

Newsome AL, et al. Isolation of an amoeba naturally harboring a distinctive Legionella species. Appl. Environ. Microbiol. 64: 1688-1693, 1998. PubMed: 9572937

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Qureshi MN, et al. Inhibition of Acanthamoeba species by Pseudomonas aeruginosa rationale for their selective exclusion in corneal ulcers and contact lens care systems. J. Clin. Microbiol. 31: 1908-1910, 1993. PubMed: 8349772

Reveiller FL, et al. Isolation of a unique membrane protein from Naegleria fowleri. J. Eukaryot. Microbiol. 48: 676-682, 2001. PubMed: 11831777

Schuster FL. Comparative effects of selected azole compounds on trophic and cystic stages of Acanthamoeba polyphaga. J. Eukaryot. Microbiol. 40: 563-569, 1993. PubMed: 8401469

Simione FP Jr., et al. The use of plastic ampoules for freeze preservation of microorganisms. Cryobiology 14: 500-502, 1977. PubMed: 891238

Visvesvara GS, Balamuth W. Comparative studies on related free-living and pathogenic amebae with special reference to Acanthamoeba. J. Protozool. 22: 245-256, 1975. PubMed: 807717

Visvesvara GS, et al. Isolation, identification, and biological characterization of Acanthamoeba polyphaga from a human eye. Am. J. Trop. Med. Hyg. 24: 784-790, 1975. PubMed: 811126

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Toney DM, Marciano-Cabral F. Resistance of Acanthamoeba species to complement lysis. J. Parasitol. 84: 338-344, 1998. PubMed: 9576508

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.
Basic Documentation