Plasmodium berghei Vincke and Lips
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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.
1. To harvest parasites, inject host with ketamine (0.10.2 ml).
2. Open chest cavity to expose heart and exsanguinate via cardiac puncture using Yaeger's anticoagulant** (see below), 1 volume anticoagulant to 4 volumes blood. Glass distilled HGla
3. Centrifuge blood for 5 mins. at 1800 rpm in 50 ml centrifuge tube.
4. Aspirate supernatant using sterile Pasteur pipet.
5. Resuspend pellet gently in remaining supernatant.
6. Slowly add 5 volumes of glycerolyte medium to 3 volumes pellet dropwise via a syringe as follows:
A. Add the first volume of glycerolyte and allow the tube to stand for 5 mins. at room temperature.
B. Add the remaining 4 volumes of glycerolyte and gently agitate.
7. 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.).
8. Plunge vials into liquid nitrogen (-196oC) the next day and store in liquid nitrogen or liquid nitrogen vapor.
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Branton MBStudies of clonal populations of NK65 Plasmodium berghei Ph.D. thesis, Univ. Illinois, 1978
Alger NE, Keshavarz-Valian H. Plasmodium berghei: the effects of suppressor factor on vaccination. Int. J. Parasitol. 14: 301-307, 1984. PubMed: 6381347