Entamoeba histolytica Schaudinn
30015 ™
- Product category
-
Protists
- Product type
-
Parasitic protozoan
- Classification
- Amoebozoa, Entamoebida
- Strain designation
- HK-9
- Type strain
- No
- Isolation source
- Colonic biopsy of adult male with amebic dysentery
- Geographical isolation
- Korea
- Applications
-
Enteric disease researchInfectious disease research
- Product format
- Frozen
- Storage conditions
-
-80°C or colder for 1 week, vapor phase of liquid nitrogen for long-term storage
Documentation
ATCC determines the biosafety level of a material based on our risk assessment as guided by the current edition of Biosafety in Microbiological and Biomedical Laboratories (BMBL), U.S. Department of Health and Human Services. It is your responsibility to understand the hazards associated with the material per your organization’s policies and procedures as well as any other applicable regulations as enforced by your local or national agencies.
ATCC highly recommends that appropriate personal protective equipment is always used when handling vials. For cultures that require storage in liquid nitrogen, it is important to note that some vials may leak when submersed in liquid nitrogen and will slowly fill with liquid nitrogen. Upon thawing, the conversion of the liquid nitrogen back to its gas phase may result in the vial exploding or blowing off its cap with dangerous force creating flying debris. Unless necessary, ATCC recommends that these cultures be stored in the vapor phase of liquid nitrogen rather than submersed in liquid nitrogen.
Required Products
These products are vital for the proper use of this item and have been confirmed as effective in supporting functionality. If you use alternative products, the quality and effectiveness of the item may be affected.
Detailed product information
General
- Specific applications
-
Enteric ResearchFood and waterborne pathogen research
Characteristics
- Comments
-
Entamoeba phylogeny
Zymodeme II
Not detected by Trichomonas vaginalis-specific PCR primers
Axenic cultivation
Use of plastic ampoules for freeze preservation
Growth responses to reducing agents
Effect of cysteine and oxygen tension on survival
Requirement for cysteine and ascorbic acid for attachment
Handling information
- Medium
- Instruction for complete medium
- ATCC Medium 2154 (Quality controlled freeze-dried lots of this medium are commercially available from ATCC)
- Temperature
-
35°C
- Atmosphere
-
Anaerobic
- Culture system
- Axenic
- Handling procedure
-
Storage and Culture Initiation
Frozen amules packed in dry ice should either be thawed immediately or stored in liquid nitrogen. If liquid nitrogen storage facilities are not available, frozen ampules may be stored at or below -70°C for approximately one week. Do not under any circumstance store frozen ampules at refrigerator freezer temperatures (generally -20°C). Storage of frozen material at this temperature will result in the death of the culture.- To thaw a frozen ampule, place in a 35°C water bath, until thawed (2 to 3 min). Immerse the ampule just sufficiently to cover the frozen material. Do not agitate the ampule.
- Immediately after thawing, aseptically transfer contents to a glass screw-capped tube containing 13 mL ATCC Medium 2154. Screw cap on tightly and incubate on a 15° horizontal slant at 35°C.
- Culture maintenance
-
- Ice culture at or near peak density for 10 min.
- Gently invert culture 20 times.
- Aseptically transfer a 0.1 mL and 0.25 mL aliquot to freshly prepared (no older than 7 to 10 days) tubes of ATCC medium 2154.
- Screw caps on tightly and incubate at a 15° horizontal slant at 35°C.
- Subculture when many trophozoites are observed (typically every 2 to 4 days). The transfer interval will depend on the quantity of the inoculum and the quality of the medium. This should be empirically determined by examining the culture on a daily basis until the growth cycle has stabilized. Do not allow the culture to overgrow. The culture crashes soon after reaching peak density.
- Reagents for cryopreservation
-
CPMB-5 Cryoprotective Solution
DMSO: 1.0 mL
2.5 M Sucrose: 0.8 mL
L-Cysteine/Ascorbic Acid Solution: 0.2 mL
CPMB-2 Basal Solution: 6.0 mL
HIBS: 2.0 mL
CPMB-2 Basal Solution
Yeast Extract: 60.0 g
K2HPO4: 1.0 g
KH2PO4: 0.6 g
NaCl: 2.0 g
Distilled water: 1.0 L
Autoclave for 15 minutes.
L-Cysteine/Ascorbic Acid Solution
L-Cysteine-HCL: 1.0 g
Acorbic Acid: 0.1 g
Distilled water: 10.0 mL
Add 9.0 mL of distilled water to a 20 mL beaker and dissolve the first two components. While stirring, adjust the pH to 7.2 with 10 M NaOH (approximately 0.7 mL). Adjust final volume to 10 mL with distilled water and filter sterilize. Solution should be used soon after preparation. Discard any unused solution.
Harvest and Preservation - Cryopreservation
-
- Harvest cells from several cultures that are in the late logarithmic to early stationary phase of growth. Place culture vessels on ice for 10 min.
- Invert tubes 20 times and centrifuge at 200 x g for 5 min.
- While cells are centrifuging, prepare the cryoprotective solution.
- Place 1.0 mL of DMSO in a 16 x 125 mm screw-capped test tube and ice until solidified.
- Add 0.8 mL of the 2.5 M Sucrose solution, remove from ice and invert until the DMSO is liquefied. Return to ice bath.
- Add 0.2 mL of the L-Cysteine/Ascorbic Acid Solution to the DMSO solution and mix.
- Add 6.0 mL of the CPMB-2 Basal Solution and mix.
- Add 2.0 mL HIBS and mix.
- Resuspend the cell pellets and pool to a final volume of approximately 10 mL with the supernatant. Make a determination of the cell density and adjust the concentration of the cells between 5 x 105/mL to 1 x 106/mL using fresh medium. If the cell concentration is below 5 x 105/mL, centrifuge the cell suspension and resuspend the pellet in a volume that will yield the desired concentration.
- After the cell concentration is adjusted, centrifuge as in step 2.
- Remove as much supernatant as possible and determine the volume removed.
- Resuspend the cell pellet with a volume of the cryoprotective solution equal to the volume of the supernatant removed. Invert the tube several times to obtain a uniform cell density.
- Dispense 0.5 mL aliquots into 1.0 mL to 2.0 mL plastic sterile cryules (special plastic vials for cryopreservation).
- Place the vials in a controlled rate freezing unit.
Use the following cooling cycle:- From room temperature cool at -10°C/min to the heat of fusion
- From the heat of fusion to -40°C, cool at -1°C/min
- At -40°C plunge into liquid nitrogen
- Store ampules in a liquid nitrogen refrigerator until needed.
- To establish a culture from the frozen state, place an ampule in a 35°C water bath, until thawed (2 to 3 min). Immerse the vial just sufficiently to cover the frozen material. Do not agitate the ampule.
- Transfer contents of thawed ampule to a 16 x 125 mm screw-capped borosilicate glass test tube containing 13 mL of ATCC medium 2154.
- Screw cap on tightly and incubate at a 15° horizontal slant at 35°C. Observe the culture daily and transfer when many trophozoites are observed.
History
- Deposited as
- Entamoeba histolytica Schaudinn
- Depositors
- LS Diamond
- Type of isolate
- Human
- Patient age
- adult
- Patient gender
- Male
- Special collection
-
NCRR Contract
Legal disclaimers
- Intended use
- This product is intended for laboratory research use only. It is not intended for any animal or human therapeutic use, any human or animal consumption, or any diagnostic use.
- Warranty
-
The product is provided 'AS IS' and the viability of ATCC® products is warranted for 30 days from the date of shipment, provided that the customer has stored and handled the product according to the information included on the product information sheet, website, and Certificate of Analysis. For living cultures, ATCC lists the media formulation and reagents that have been found to be effective for the product. While other unspecified media and reagents may also produce satisfactory results, a change in the ATCC and/or depositor-recommended protocols may affect the recovery, growth, and/or function of the product. If an alternative medium formulation or reagent is used, the ATCC warranty for viability is no longer valid. Except as expressly set forth herein, no other warranties of any kind are provided, express or implied, including, but not limited to, any implied warranties of merchantability, fitness for a particular purpose, manufacture according to cGMP standards, typicality, safety, accuracy, and/or noninfringement.
- Disclaimers
-
This product is intended for laboratory research use only. It is not intended for any animal or human therapeutic use, any human or animal consumption, or any diagnostic use. Any proposed commercial use is prohibited without a license from ATCC.
While ATCC uses reasonable efforts to include accurate and up-to-date information on this product sheet, ATCC makes no warranties or representations as to its accuracy. Citations from scientific literature and patents are provided for informational purposes only. ATCC does not warrant that such information has been confirmed to be accurate or complete and the customer bears the sole responsibility of confirming the accuracy and completeness of any such information.
This product is sent on the condition that the customer is responsible for and assumes all risk and responsibility in connection with the receipt, handling, storage, disposal, and use of the ATCC product including without limitation taking all appropriate safety and handling precautions to minimize health or environmental risk. As a condition of receiving the material, the customer agrees that any activity undertaken with the ATCC product and any progeny or modifications will be conducted in compliance with all applicable laws, regulations, and guidelines. This product is provided 'AS IS' with no representations or warranties whatsoever except as expressly set forth herein and in no event shall ATCC, its parents, subsidiaries, directors, officers, agents, employees, assigns, successors, and affiliates be liable for indirect, special, incidental, or consequential damages of any kind in connection with or arising out of the customer's use of the product. While reasonable effort is made to ensure authenticity and reliability of materials on deposit, ATCC is not liable for damages arising from the misidentification or misrepresentation of such materials.
Please see the material transfer agreement (MTA) for further details regarding the use of this product. The MTA is available at www.atcc.org.
Permits & Restrictions
Import Permit for the State of Hawaii
If shipping to the U.S. state of Hawaii, you must provide either an import permit or documentation stating that an import permit is not required. We cannot ship this item until we receive this documentation. Contact the Hawaii Department of Agriculture (HDOA), Plant Industry Division, Plant Quarantine Branch to determine if an import permit is required.
Frequently Asked Questions
References
Curated Citations
Charoenlarp P, et al. Carbohydrate utilization by Entamoeba histolytica. Exp. Parasitol. 23: 205-211, 1968. PubMed: 4302387
Clark CG, Diamond LS. Intraspecific variation and phylogenetic relationships in the genus Entamoeba as revealed by riboprinting. J. Eukaryot. Microbiol. 44: 142-154, 1997. PubMed: 9109261
Diamond LS. Axenic cultivation of Entamoeba histolytica. Science 134: 336-337, 1961. PubMed: 13722605
Diamond LS. Techniques of axenic cultivation of Entamoeba histolytica Schaudinn, 1903 and E. histolytica-like amebae. J. Parasitol. 54: 1047-1056, 1968. PubMed: 4319346
Garcia LS, et al. Comparison of indirect fluorescent-antibody amoebic serology with counterimmunoelectrophoresis and indirect hemagglutination amoebic serologies. J. Clin. Microbiol. 15: 603-605, 1982. PubMed: 6279694
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