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Product Name:
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RH FLUC
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Manufacturer:
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BEI Resources
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Taxonomy:
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Protozoa Classification: Apicomplexa, Toxoplasma
Species: Toxoplasma gondii
Strain: RH FLUC
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Additional Information:
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T. gondii is an obligate intracellular protozoan parasite of the phylum Apicomplexa that is the causal agent of toxoplasmosis. T. gondii has a highly unusual, clonal population structure comprised of three widespread genotypes referred to as type I (highly virulent), type II (nonvirulent), and type III (associated with animal infections), which account for more than 95% of strains isolated in North America and Europe.4,5,6,7 Isolates from South America exhibit greater genetic diversity. Phylogenetic analyses of T. gondii intron sequences have identified eleven separate haplogroups, with striking geographic separation between North America, Europe and South America.8
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Material Provided:
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Each vial of NR-59752 contains approximately 0.5 mL of culture in cryopreservative [7.5% dimethylsulfoxide (DMSO)]. Please refer to Appendix I for cryopreservation instructions.
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Packing/Storage:
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NR-59752 was packaged aseptically in cryovials and is provided frozen on dry ice. The product should be stored at -130°C or colder, preferably in the vapor phase of a liquid nitrogen freezer. If liquid nitrogen storage facilities are not available, frozen cryovials may be stored at -70°C or colder for approximately one week.
Note: Do not under any circumstances store vials at temperatures warmer than -70°C. Storage under these conditions will result in the death of the culture.
To ensure the highest level of viability, the culture should be initiated immediately upon receipt. Any warming of the product during shipping and transfer must be avoided, as this will adversely affect the viability of the product. For transfer between freezers and for shipping, the product may be placed on dry ice for brief periods, although use of a portable liquid nitrogen carrier is preferred. Please read the following recommendations prior to using this material.
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Growth Conditions:
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Media:
Dulbecco's Minimal Essential Medium (DMEM) supplemented with 10% (v/v) heat-inactivated fetal bovine serum (HIFBS)
Human foreskin fibroblast cells (ATCC® CRL-1634™)
Incubation:
Temperature: 37°C
Atmosphere: Aerobic with 5% CO2
Propagation:
1. To establish a culture from the frozen state, place a vial in a 35°C to 37°C water bath. Thawing time is approximately 2 to 3 minutes. Do not agitate the vial. Do not leave the vial in the water bath after it is thawed.
2. Immediately after thawing, aseptically transfer the contents to a culture flask containing a fresh monolayer of human foreskin fibroblast cells (ATCC® CRL-1634™) and 10 mL of DMEM containing 10% (v/v) HIFBS.
3. Outgas the flask for 10 seconds with a 95% air, 5% CO2 gas mixture.
4. Incubate at 37°C in an aerobic atmosphere with 5% CO2 with the caps screwed on tightly. Observe the culture daily under an inverted microscope for the presence of parasitophorous vacuoles.
Maintenance:
1. Remove the medium from a fresh confluent monolayer of human foreskin fibroblast cells in a tissue culture flask and replace it with 10 mL medium containing 10% (v/v) HIFBS.
2. Remove the medium from the Toxoplasma culture when approximately 50% of the human foreskin fibroblast cell monolayer has lysed. Centrifuge the parasites that had been released into the medium at 1300 × g for 10 minutes.
3. Remove the supernatant and resuspend the cell pellet in a small volume (0.5 mL to 1 mL) of DMEM containing 10% (v/v) HIFBS or phosphate buffered saline (PBS). Transfer the resuspended pellet to the fresh flask of human foreskin fibroblast cells prepared in step 1 above. Follow steps 3 and 4 in Propagation.
Please refer to Appendix I for cryopreservation instructions.
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Disclaimers:
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You are authorized to use this product for research use only. It is not intended for human use. Use of this product is subject to the terms and conditions of the BEI Resources Material Transfer Agreement (MTA). The MTA is available on our Web site at www.beiresources.org. While BEI Resources uses reasonable efforts to include accurate and up-to-date information on this product sheet, neither ATCC® nor the U.S. Government makes any warranties or representations as to its accuracy. Citations from scientific literature and patents are provided for informational purposes only. Neither ATCC® nor the U.S. Government warrants that such information has been confirmed to be accurate. This product is sent with the condition that you are responsible for its safe storage, handling, use and disposal. ATCC® and the U.S. Government are not liable for any damages or injuries arising from receipt and/ or use of this product. While reasonable effort is made to ensure authenticity and reliability of materials on deposit, the U.S. Government, ATCC®, their suppliers and contributors to BEI Resources are not liable for damages arising from the misidentification or misrepresentation of products. |
References:
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1. Sibley, L. D., Personal Communication.
2. Radke, J. B., et al. “Bicyclic Azetidines Target Acute and Chronic Stages of Toxoplasma gondii by Inhibiting Parasite Phenylalanyl t-RNA Synthase.” Nat. Commun. 13 (2022): 459. PubMed: 35075105.
3. Huynh, M. and V. B. Carruthers. “Tagging of Endogenous Genes in a Toxoplasma gondii Strain lacking Ku80.” Eukaryot. Cell. 8 (2009): 530-539. PubMed: 19218426.
4. Howe, D. K. and L. D. Sibley. “Toxoplasma gondii Comprises Three Clonal Lineages: Correlation of Parasite Genotype with Human Disease.” J. Infect. Dis. 172 (1995): 1561-1566. PubMed: 7594717.
5. Sibley, L. D. and J. C. Boothroyd. “Virulent Strains of Toxoplasma gondii Comprise a Single Clonal Lineage.” Nature 359 (1992): 82-85. PubMed: 1355855.
6. Khan, A., et al. “Composite Genome Map and Recombination Parameters Derived from Three Archetypal Lineages of Toxoplasma gondii.” Nucleic Acids Res. 33 (2005): 2980-2992. PubMed: 15911631.
7. Sibley, L. D., et al. “Generation of a Restriction Fragment Length Polymorphism Linkage Map for Toxoplasma gondii.” Genetics 132 (1992): 1003-1015. PubMed: 1360931.
8. Khan, A., et al. “Recent Transcontinental Sweep of Toxoplasma gondii Driven by a Single Monomorphic Chromosome.” Proc. Natl. Acad. Sci. USA 104 (2007): 14872-14877. PubMed: 17804804.
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Citation:
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Acknowledgment for publications should read “The following reagent was obtained through BEI Resources, NIAID, NIH: Toxoplasma gondii, Strain RH FLUC, NR-59752.”
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Biosafety Level:
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2
Appropriate safety procedures should always be used with this material. Laboratory safety is discussed in the following publication: U.S. Department of Health and Human Services, Public Health Service, Centers for Disease Control and Prevention, and National Institutes of Health. Biosafety in Microbiological and Biomedical Laboratories (BMBL). Current Edition. Washington, DC: U.S. Government Printing Office.
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