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1. Harvest cells from a culture that is at or near peak density. To detach cells from the wall of the culture tubes place on ice for 10 minutes.  Invert tubes several times until the majority of the cells are in suspension.  Centrifuge tubes at 800 x g for 5 minutes. 
2. Adjust the concentration of cells to 2 x 10⁷/mL in fresh medium.
3. Before centrifuging, prepare a 24% (v/v) solution of sterile DMSO in fresh medium containing 8% (w/v) sucrose.  The solution is prepared as follows:
   1. Add 1.05 g sucrose to 10 mL of fresh medium and filter sterilize through a 0.2 mm filter;
   2. Add 2.4 mL of DMSO to an ice cold 20 x 150 mm screw-capped test tube;
   3. Place the tube on ice and allow the DMSO to solidify (~5 min) and then add 7.6 mL of ice cold medium prepared in step 3a.  The final concentration will be 24% (v/v) DMSO and 8% (w/v) sucrose;
   4. Invert several times to dissolve the DMSO;
   5. Allow to warm to room temperature.
4. Mix the cell preparation and the cryoprotective agent, prepared in step 3, in equal portions. Thus, the final concentration will equal 12% (v/v) DMSO + 4% sucrose (w/v) and 10⁷ cells/mL. 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 should be stored in either the vapor or liquid phase of a nitrogen refrigerator. Frozen preparations stored below -130°C are stabile indefinitely. Those stored at temperatures above -130°C are progressively less stabile as the storage temperature is elevated.
8. To establish a culture from the frozen state place an ampule in a water bath set at 35°C. Immerse the vial just to a level just above the surface of the frozen material. Do not agitate the vial.
9. After thawing, do not leave in the water bath; rather, immediately   aseptically remove the contents of the ampule and inoculate a 16 x 125 mm screw-capped test tube containing 13 mL ATCC Medium 2695.
10. Incubate the culture on a 15° horizontal slant at 35°C.

Nash TE, et al. Restriction-endonuclease analysis of DNA from 15 Giardia isolates obtained from humans and animals. J. Infect. Dis. 152: 64-73, 1985. PubMed: 2409186

Kabnick KS, Peattie DA. In situ analyses reveal that the two nuclei of Giardia lamblia are equivalent. J. Cell Sci. 95: 353-360, 1990. PubMed: 2384520

Weinbach EC, et al. Respiratory metabolism of Giardia lamblia. J. Parasitol. 66: 347-350, 1980. PubMed: 6248623

Isaac-Renton JL, et al. Characterization of Giardia duodenalis isolates from a waterborne outbreak. J. Infect. Dis. 167: 431-440, 1993. PubMed: 8421176

Gillin FD, Reiner DS. Attachment of the flagellate Giardia lamblia: Role of reducing agents, serum, temperature, and ionic composition. Mol. Cell. Biol. 2: 369-377, 1982. PubMed: 7110136

Nash TE, et al. Excretory-secretory products of Giardia lamblia. J. Immunol. 131: 2004-2010, 1983. PubMed: 6619547

Weinbach EC, et al. Binding of tricyclic antidepressant drugs to trophozoites of Giardia lamblia. Comp. Biochem. Physiol. 102: 391-396, 1992. PubMed: 1360349

Monis PT, et al. Molecular genetic analysis of Giardia intestinalis isolates at the glutamate dehydrogenase locus. Parasitology 112: 1-12, 1996. PubMed: 8587793

Munoz ML, et al. Giardia lamblia: Detection and characterization of calmodulin. Exp. Parasitol. 63: 42-48, 1987. PubMed: 3026834

McCabe RE, et al. In vitro model of attachment of Giardia intestinalis trophozoites to EIC-6 cells, an intestinal cell line. Antimicrob. Agents Chemother. 35: 29-35, 1991. PubMed: 1901700

Meloni BP, et al. Critical comparison of Giardia duodenalis from Australia and Switzerland using isoenzyme electrophoresis. Acta Trop. 50: 115-124, 1992. PubMed: 1685867

Gillin FD, et al. Human milk kills parasitic intestinal protozoa. Science 221: 1290-1292, 1983. PubMed: 6310751

Jimenez BM, et al. Inhibition of uridine phosphorylase from Giardia lamblia by pyrimidine analogs. Biochem. Pharmacol. 38: 3785-3789, 1989. PubMed: 2597172

Wieder SC, et al. Mass cultivation of Giardia lamblia in a serum-free medium. J. Parasitol. 69: 1181-1182, 1983. PubMed: 6674471

Gillin FD, Diamond LS. Clonal growth of Giardia lamblia trophozoites in a semisolid agarose medium. J. Parasitol. 66: 350-352, 1980. PubMed: 7391877

Bertram MA, et al. A comparison of isozymes of five axenic Giardia isolates. J. Parasitol. 69: 793-801, 1983. PubMed: 6672161

Binz N, et al. A simple method for cloning Giardia duodenalis from cultures and fecal samples. J. Parasitol. 77: 627-631, 1991. PubMed: 1865272

Gillin FD, et al. Inhibition of growth of Giardia lamblia by difluoromethylornithine, a specific inhibitor of polyamine biosynthesis. J. Protozool. 31: 161-163, 1984. PubMed: 6330350

Peattie DA, et al. Ultrastructural localization of giardins to the edges of disk microribbons of Giarida lamblia and the nucleotide and deduced protein sequence of alpha giardin. J. Cell Biol. 109: 2323-2335, 1989. PubMed: 2808530

Finch GR, et al. Comparison of Giardia lamblia and Giardia muris cyst inactivation by ozone. Appl. Environ. Microbiol. 59: 3674-3680, 1993. PubMed: 8285675

Swarbrick A, et al. Nucleotide variaation in the cytidine triphosphate synthetase gene of Giardia duodenalis. J. Eukaryot. Microbiol. 44: 531-534, 1997. PubMed: 9435124

Madico G, et al. Diagnosis of Trichomonas vaginalis infection by PCR using vaginal swab samples. J. Clin. Microbiol. 36: 3205-3210, 1998. PubMed: 9774566

Ey PL, Darby JM. VSP417-6, a variant-specific surface protein encoded at a sixth locus within the vsp417 gene subfamily of Giardia intestinalis. Int. J. Parasitol. 32: 425-436, 2002. PubMed: 11849639

Karanis P, Ey PL. Characterization of axenic isolates of Giardia intestinalis established from humans and animals in Germany. Parasitol. Res. 84: 442-449, 1998. PubMed: 9660132

Miller RL, et al. Identification of Giardia lamblia Isolates Susceptible and Resistant to Infection by the Double-Stranded RNA Virus. Exp. Parasitol. 66: 118-123, 1988. PubMed: 3366209