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Plasmodium falciparum Welch

50110

Product category
Protists
Product type
Parasitic protozoan
Classification
KINGDOM: Protozoa
Strain designation
HB-2
Type strain
No
Isolation source
Derived from existing strain
Applications
Infectious disease research
Vector-borne disease research
Product format
Frozen
Storage conditions
-80°C or colder for 1 week, vapor phase of liquid nitrogen for long-term storage
Mission Collection Item
This is a Mission Collection Item

Documentation

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Biosafety Icon BSL 2

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.

Detailed product information

General

Specific applications
Emerging infectious disease research
Vector borne research

Characteristics

Comments
This is a non-gametocyte-forming strain.

Handling information

Medium
Instruction for complete medium
ATCC Medium 2196 and type O blood
Temperature
37°C
Incubation
In vitro culture in human erythrocytes
Handling procedure

Storage and Culture Initiation

Frozen ampules packed in dry ice should either be thawed immediately or stored in liquid nitrogen.  If liquid nitrogen storage facilities are not available, frozen ampoules 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.

The following directions for recovery from the frozen state must be carefully followed if a culture is to be successfully established.

  1. Place the frozen vial in a 37°C water bath until mixture is completely thawed.
  2. Aseptically transfer the contents to a 50 ml sterile conical tube.
  3. Slowly add 1 volume (0.1 ml) 12% Sodium Chloride solution dropwise via a 1ml syringe to 5 volumes sample (0.5 ml) and agitate continuously.
  4. Allow the mixture to stand for 5 mins. at room temperature.
  5. Slowly add 5 ml 1.8% Sodium Chloride dropwise via a larger syringe and allow to stand at room temperature for 2 mins.
  6. Add 5 ml of 0.9% Sodium Chloride / 0.2% Glucose solution as in step 5.
  7. Centrifuge for 5 min. at 1500rpm, remove supernatant.
  8. Wash pellet in 20 ml incomplete medium.
  9. Centrifuge for 5 min at 1500 rpm, remove supernatant.
  10. Resuspend pellet in 8ml complete medium in tissue culture flask and gently aerate culture with gas mixture of 5% CO2, 5% O2 and 90% N2 using a sterile, cotton plugged Pasteur pipet.
  11. Smear if required (see below).


* Medium Formulation:

To a 500 ml bottle of RPMI-1640 (with NaHCO3, without L-glutamine), aseptically add 18.75ml HEPES (final = 37.5mM), 5ml L-glutamine, 0.25ml Gentamicin, 5ml 20% Glucose (final = 20mM), 3ml 1M Sodium Hydroxide (pH > 7), 5ml Hypoxanthine 100x .  Mix thoroughly and remove 100ml from the bottle to make incomplete medium; then add 50ml of heat inactivated human serum (any type) to the remainder to make complete medium (10% serum). Store at 4°C.

NOTE: Outdated blood and serum cannot be used to cultivate this strain. Use only fresh blood and serum.

Culture maintenance
Changing of the culture medium every 24 hours is required for a malaria-infected erythrocyte culture.

  1. Remove flask with infected culture from 37°C incubator and place onto flask warmer in biological safety hood.
  2. Carefully aspirate the medium with a sterile unplugged Pasteur pipet attached to a vacuum line. Remove as much fluid as possible without taking the cells.
  3. Add 25 ml of sterile warm (37°C) complete medium to the flask, gently mix and aerate, then quickly tighten cap of the flask and place in 37°C incubator until next medium change.


Making a Blood Smear:

  1. Aseptically transfer 0.5–1ml of mixed culture with a sterile pipet into an eppendorf tube.
  2. Spin down the eppendorf tube at high speed and aspirate the supernatant.
  3. Mix the pellet and place a drop of the suspension on a glass slide. Spread the drop into a thin film with the edge of another glass slide. Air dry for 3 mins. at room temperature.
  4. Fix air-dried blood film by rinsing with methyl alcohol. Air dry for 3 mins. at room temperature.
  5. Stain blood films in 5% Giemsa solution for 15 mins. Rinse with distilled water, air dry.
  6. Observe using light microscopy at 100X magnification to determine parasitemia of culture.
Cryopreservation

Only young cells (rings) can be frozen in glycerolyte medium** because their membranes are more robust.

  1. Centrifuge ring stage culture for 5 min at 1800rpm in 50 ml centrifuge tube.
  2. Aspirate supernatant using sterile Pasteur pipet.
  3. Resuspend pellet gently in remaining supernatant.
  4. Slowly add 5 volumes of glycerolyte medium (see below) to 3 volumes pellet dropwise via a syringe as follows:
    1. Add the first volume of glycerolyte and allow the tube to stand for 5 mins. at room temperature.
    2. Add the remaining 4 volumes of glycerolyte and gently agitate.
  5. Aliquot mixture into Nunc screwtop freezing vials and store at -80°C overnight.
  6. Plunge vials into liquid nitrogen (-196°C) the next day and store in liquid nitrogen or liquid nitrogen vapor.

** To formulate glycerolyte medium, combine the following with distilled water to 100 ml:  57.00g glycerol, 1.60g sodium lactate (C3H5NaO3), 30.00mg potassium chloride (KCl), 1.38g sodium dihydrogen phosphate (NaH2PO4). Mix well and adjust pH to 6.8 using concentrated NaOH and/or HCl. Autoclave to sterilize, and store at 4°C.

History

Deposited as
Plasmodium falciparum Welch
Depositors
W Trager
Year of origin
1981

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.

MORE INFORMATION ABOUT PERMITS AND RESTRICTIONS

References

Curated Citations

Bhasin VK, Trager W. Gametocyte-forming and non-gametocyte-forming clones of Plasmodium falciparum. Am. J. Trop. Med. Hyg. 33: 534-537, 1984. PubMed: 6383092

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