Crithidia ricardoi Sibajev et al. (ATCC® 50523)

Depositor: A Sibajev  /  Biosafety Level: 1

Permits and Restrictions

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Biosafety Level 1
Isolation
mosquito
Isolation date: March 7, 1991
Product Format frozen
Type Strain no
Medium Medium 1034: Modified PYNFH medium (Available from ATCC as ATCC cat. no. 327-X)
Growth Conditions
Temperature: 25.0°C
Duration: axenic
Protocol: ATCCNO: 11745 SPEC: See general instructions for thawing and storage of frozen material before proceeding. Add thawed contents to a single 16 x 125 mm glass screw-capped test tube of the appropriate medium. Incubate the culture vertically with the cap screwed on tightly. It is essential to establish cultures initially in small volumes. Once established, the culture can be scaled up to larger volumes. Vigorously agitate the culture and aseptically transfer 0.1 ml of culture to a fresh tube of medium weekly.
Subcultivation
Protocol: ATCCNO: 11745 SPEC: See general instructions for thawing and storage of frozen material before proceeding. Add thawed contents to a single 16 x 125 mm glass screw-capped test tube of the appropriate medium. Incubate the culture vertically with the cap screwed on tightly. It is essential to establish cultures initially in small volumes. Once established, the culture can be scaled up to larger volumes. Vigorously agitate the culture and aseptically transfer 0.1 ml of culture to a fresh tube of medium weekly.
Cryopreservation

1.   Prepare a 10% (v/v) sterile DMSO solution in fresh ATCC Medium 355. 

2.   Transfer a culture at peak density to centrifuge tubes and centrifuge at 525 x g for 5 minutes.

3.   Remove the supernatant and resuspend the cells in ATCC medium 355 to a concentration of 2 x 106 to 2 x 107 cells/ml.

4.   Mix the cell preparation and the DMSO in equal portions. Thus, the final concentration will be between 106 and 107 cells/ml and 5% (v/v) DMSO.

5.   Distribute the cell suspension in 0.5 ml aliquots into 1.0-2.0 ml sterile plastic screw-capped cryules (special plastic vials for cryopreservation).  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.

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 are stored in either the vapor or liquid phase of a nitrogen freezer.

8.   To establish a culture from the frozen state place an ampule in a water bath set at 35°C (2-3 min). Immerse the vial just sufficient to cover the frozen material. Do not agitate the vial.

9.   Immediately after thawing, aseptically remove the contents of the ampule and inoculate into 5 ml of fresh ATCC medium 355 in a 16 x 125 mm screw-capped test tube. Incubate upright at 25°C with caps screwed on tightly.

Name of Depositor A Sibajev
Year of Origin March 7, 1991
References

Sibajev A, et al. Crithidia ricardoi sp. n. a new species of Trypanosomatidae isolated from Culex saltanensis Dyar, 1928 (Diptera; Culicidae). Mem. Inst. Oswaldo Cruz Rio J. 88: 541-545, 1993.

digestive tract of mosquito, Culex saltanensis, Rio de Janeiro, Brazil

Notice: Necessary PermitsPermits

These permits may be required for shipping this product:

  • Customers located in the state of Hawaii will need to contact the Hawaii Department of Agriculture to determine if an Import Permit is required. A copy of the permit or documentation that a permit is not required must be sent to ATCC in advance of shipment.
Basic Documentation