Biosafety classification is based on U.S. Public Health Service Guidelines, it is the responsibility of the customer to ensure that their facilities comply with biosafety regulations for their own country.
Isolation
Mourning dove, Nebraska, 1937
Product Format
frozen
Storage Conditions
Frozen: -70°C or colder Freeze-Dried: 2°C to 8°C Live Culture: See Protocols Section
Type Strain
no
Growth Conditions
In-vivo cultivation in pigeons (age 3-4 d.)
Cryopreservation
Reagents Glycerolyte medium
Glycerol, 57.00 g
NaC3H5O3 (Sodium lactate), 1.60 g
KCl, 30.00 mg
NaH2PO4, 1.38 g
Glass distilled water, 100.00 mL
Mix well and adjust pH to 6.8 using concentrated NaOH and/or HCl. Autoclave and store at 4°C.
Yaeger's anticoagulant
Sodium citrate, 1.33 g
Citric acid, 0.47 g
Dextrose, 3.00 g
Sodium heparin, 0.20 g
Glass distilled water, 100.00 mL
Filter sterilize.
Harvest and Preservation
Only young cells (rings) can be frozen in glycerolyte medium* because their membranes are more robust.
To harvest parasites, inject host with ketamine (0.1–0.2 mL).
Exsanguinate via the wing vein using a sterile syringe and Yaeger's anticoagulant, 1 volume anticoagulant to 4 volumes blood.
Centrifuge blood for 5 mins. at 1800 rpm in 50 mL centrifuge tube.
Aspirate supernatant using sterile Pasteur pipet.
Resuspend pellet gently in remaining supernatant.
Slowly add 5 volumes of glycerolyte medium to 3 volumes pellet dropwise via a syringe as follows:
Add the first volume of glycerolyte and allow the tube to stand for 5 mins. at room temperature.
Add the remaining 4 volumes of glycerolyte and gently agitate.
Aliquot mixture into Nunc screw-capped freezing vials and place in a Nalgene 1°C cooling apparatus. Place the apparatus at -80°C overnight and then plunge ampules into liquid nitrogen. (The cooling rate in this apparatus is approximately -1°C/min.).
Plunge vials into liquid nitrogen (-196°C) the next day and store in liquid nitrogen or liquid nitrogen vapor.
Name of Depositor
RG Yaeger
Chain of Custody
ATCC <-- RG Yaeger <-- GH Ball <-- C Huff <-- GR Coatney
