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Purification of white spot syndrome virus by iodixanol density gradient centrifugation
Dantas-Lima, J.J.; Corteel, M.; Cornelissen, M.; Bossier, P.; Sorgeloos, P.; Nauwynck, H.J. (2013). Purification of white spot syndrome virus by iodixanol density gradient centrifugation. J. Fish Dis. 36(10): 841-851. https://dx.doi.org/10.1111/jfd.12082
In: Journal of Fish Diseases. Blackwell Science: Oxford; London; Edinburgh; Boston; Melbourne. ISSN 0140-7775; e-ISSN 1365-2761
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| Trefwoord |
Penaeus vannamei Boone, 1931 [WoRMS]
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| Author keywords |
iodixanol; Penaeus vannamei; white spot syndrome virus buoyant density;white spot syndrome virus infectivity; white spot syndrome viruspurification |
| Auteurs | | Top |
- Dantas-Lima, J.J.
- Corteel, M.
- Cornelissen, M.
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- Bossier, P.
- Sorgeloos, P.
- Nauwynck, H.J.
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| Abstract |
Up to now, only a few brief procedures for purifying white spot syndrome virus (WSSV) have been described. They were mainly based on sucrose, NaBr and CsCl density gradient centrifugation. This work describes for the first time the purification of WSSV through iodixanol density gradients, using virus isolated from infected tissues and haemolymph of Penaeus vannamei (Boone). The purification from tissues included a concentration step by centrifugation (2.5h at 60000g) onto a 50% iodixanol cushion and a purification step by centrifugation (3h at 80000g) through a discontinuous iodixanol gradient (phosphate-buffered saline, 5%, 10%, 15% and 20%). The purification from infected haemolymph enclosed a dialysis step with a membrane of 1000 kDa (18h) and a purification step through the earlier iodixanol gradient. The gradients were collected in fractions and analysed. The number of particles, infectivity titre (in vivo), total protein and viral protein content were evaluated. The purification from infected tissues gave WSSV suspensions with a very high infectivity and an acceptable purity, while virus purified from haemolymph had a high infectivity and a very high purity. Additionally, it was observed that WSSV has an unusually low buoyant density and that it is very sensitive to high external pressures. |
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