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1. Harvest cells from a culture that is at or near peak density by centrifugation at 600 x g for 5 min. Pool the cell pellets into a single tube.
2. Adjust the concentration of cells to 2.0 x 10⁶/mL.  If the concentration is too low, centrifuge at 600 x g for 5 minutes and resuspend the cell pellet with a volume of supernatant to yield the desired concentration.
3. Prepare a 15% (v/v) sterile DMSO solution in ATCC medium 1034 as follows:
   1. Add the required volume of DMSO to a glass screw-capped test tube and place on ice. Allow the DMSO to solidify. 
   2. Add the required volume of refrigerated ATCC medium 1034.  Dissolve the DMSO by inverting several times.  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 10⁶ and 7.5% (v/v) DMSO. The time from the mixing of the cell preparation and DMSO stock solution to the start of the freezing process should be no less than 15 min and no longer than 60 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 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.
7. The frozen preparations are stored in either the vapor or liquid phase of a nitrogen refrigerator.
8. To establish a culture from the frozen state place an ampule in a water bath set at 35°C. Immerse the vial just sufficiently to cover only the frozen material. Do not agitate the vial.
9. Immediately after thawing, do not leave in the water bath, aseptically remove the contents of the ampule and inoculate into a fresh tube or flask of ATCC medium 1034.
10. Incubate at 25°C with the cap screwed on tightly (incubate a test tube on a 15° horizontal slant).

abu Kwaik Y, et al. Protein expression by the protozoan Hartmannella vermiformis upon contact with its bacterial parasite Legionella pneumophila. Infect. Immun. 62: 1860-1866, 1994. PubMed: 8168950

Abu Kwaik Y, et al. Signal transduction in the protozoan host hartmannella vermiformis upon attachment and invasion by legionella micdadei. Appl. Environ. Microbiol. 64: 3134-3139, 1998. PubMed: 9726850

Aragon V, et al. Legionella pneumophila major acid phosphatase and its role in intracellular infection. Infect. Immun. 69: 177-185, 2001. PubMed: 11119504

Aragon V, et al. Secreted enzymatic activities of wild-type and pilD-deficient Legionella pneumophila. Infect. Immun. 68: 1855-1863, 2000. PubMed: 10722574

Brieland J, et al. Coinoculation with Hartmannella vermiformis enhances replicative Legionella pneumophila lung infection in a murine model of Legionnaires' disease. Infect. Immun. 64: 2449-2456, 1996. PubMed: 8698466

Brieland JK, et al. Host pathogen interactions in Legionnaires' disease: lessons learned from a murine animal model. Immunopharmacology 48: 249-252, 2000. PubMed: 10960664

Brieland J, et al. Intrapulmonary Hartmannella vermiformis: a potential niche for Legionella pneumophila replication in a murine model of legionellosis. Infect. Immun. 65: 4892-4896, 1997. PubMed: 9353084

Brieland JK, et al. The role of Legionella pneumophila-infected Hartmannella vermiformis as an infectious particle in a murine model of Legionnaire's disease. Infect. Immun. 65: 5330-5333, 1997. PubMed: 9393834

Cianciotto NP, Fields BS. Legionella pneumophila mip gene potentiates intracellular infection of protozoa and human macrophages. Proc. Natl. Acad. Sci. USA 89: 5188-5191, 1992. PubMed: 1594630

Flieger A, et al. Characterization of the Gene Encoding the Major Secreted Lysophospholipase A of Legionella pneumophila and Its Role in Detoxification of Lysophosphatidylcholine . Infect. Immun. 70: 6094-6106, 2002. PubMed: 12379686

Fields BS, et al. Legionella and protozoa: interaction of a pathogen and its natural host. In: Fields BS, et al., Legionella: current status and emerging perspectives. Washington DC, American Society for Microbiology, pp. 129-136, 1993

Fields BS, et al. Virulence of a Legionella anisa strain associated with Pontiac fever: an evaluation using protozoan, cell culture, and guinea pig models. Infect. Immun. 58: 3139-3142, 1990. PubMed: 2117580

Gao L-Y, et al. Heterogeneity in intracellular replication and cytopathogenicity of Legionella pneumophila and Legionella micdadei in mammalian and protozoan cells. Microb. Pathog. 27: 273-287, 1999. PubMed: 10545255

Harb OS, et al. Heterogeneity in the attachment and uptake mechanisms of the Legionnaires' disease bacterium, Legionella pneumophila, by protozoan hosts. Appl. Environ. Microbiol. 64: 126-132, 1998. PubMed: 9435069

Harb OS, Abu Kwaik Y. Identification of the aspartate-beta-semialdehyde dehydrogenase gene of Legionella pneumophila and characterization of a null mutant. Infect. Immun. 66: 1898-1903, 1998. PubMed: 9573067

King CH, et al. Effects of cytochalasin D and methylamine on intracellular growth of Legionella pneumophila in amoebae and human monocyte-like cells. Infect. Immun. 59: 758-763, 1991. PubMed: 1997428

Kwaik YA. The phagosome containing Legionella pneumophila within the protozoan Hartmannella vermiformis is surrounded by the rough endoplasmic reticulum. Appl. Environ. Microbiol. 62: 2022-2028, 1996. PubMed: 8787400

O'Connell WA, et al. A Legionella pneumophila gene that promotes hemin binding. Infect. Immun. 64: 842-848, 1996. PubMed: 8641790

Polesky AH, et al. Identification of Legionella pneumophila genes important for infection of amoebas by signature-tagged mutagenesis. Infect. Immun. 69: 977-987, 2001. PubMed: 11159993

Pope CD, et al. Legionella pneumophila mutants that are defective for iron acquisition and assimilation and intracellular infection. Infect. Immun. 64: 629-636, 1996. PubMed: 8550218

Rossier O, Cianciotto NP. Type II protein secretion is a subset of the PilD-dependent processes that facilitate intracellular infection by Legionella pneumophila. Infect. Immun. 69: 2092-2098, 2001. PubMed: 11254562

Venkataraman C, et al. Identification of putative cytoskeletal protein homologues in the protozoan host Hartmannella vermiformis as substrates for induced tyrosine phosphatase activity upon attachment to the Legionnaires' disease bacterium, Legionella pneumophila. J. Exp. Med. 188: 505-514, 1998. PubMed: 9687528

Wadowsky RM, et al. Gentamicin-containing peptone-yeast extract medium for cocultivation of Hartmannella veriformis ATCC 50256 and virulent strains of Legionella pneumophila. Appl. Environ. Microbiol. 61: 4464-4467, 1995.

Robey M, Cianciotto NP. Legionella pneumophila feoAB promotes ferrous iron uptake and intracellular infection. Infect. Immun. 70: 5659-5669, 2002. PubMed: 12228295

Robey M, et al. Identification of Legionella pneumophila rcp, a pagP-like gene that confers resistance to cationic antimicrobial peptides and promotes intracellular infection. Infect. Immun. 69: 4276-4286, 2001. PubMed: 11401964

Stone BJ, et al. The Legionella pneumophila prp locus; required during infection of macrophages and amoebae. Microb. Pathog. 27: 369-376, 1999. PubMed: 10588909

Viswanathan VK, et al. The cytochrome c maturation locus of Legionella pneumophila promotes iron assimilation and intracellular infection and contains a strain-specific insertion sequence element. Infect. Immun. 70: 1842-1852, 2002. PubMed: 11895946

Weekers PHH, et al. Sequence variations in small-subunit ribosomal RNAs of Hartmannella vermiformis and their phylogenetic implications. Mol. Biol. Evol. 11: 684-690, 1994. PubMed: 8078407

Venkataraman C, Abu Kwaik Y. Signal transduction in the protozoan host Hartmannella vermiformis upon attachment to Legionella pneumophila. Microbes Infect. 2: 867-875, 2000. PubMed: 10962269