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Iran J Virol 2019, 13(1): 35-40 Back to browse issues page
Purification of Human Reovirus in Monolayer of L-929 Cells
Mojtaba Hamidi-Fard , Angila Ataei-Pirkooh , Mohammadreza Aghasadeghi , Reihaneh Kazemi *
Hepatitis & AIDS department, Pasteur institute of Iran, Tehran, Iran.
Abstract:   (1513 Views)
Background and Aims: The  ability  of  the  reovirus  to  destroy  various  cancer  cells  in  vitro  and,  the  success  of  the  virus  in  conducting  clinical  trial  phases  of  cancer  therapy  has  attracted  the  attention  of  researchers  toward  the  virus.  One  of  the  main  needs  for  the  investigation  of  the  viral  effects  and  the  virus-host  cell  interaction  is  preparation  of  the  purified  virus.  Most  of  the  protocols  have  been  based  on  the  use  of  suspended  cell  culture  equipment  that  normally  does  not  exist  in  the  research  laboratories.  We  optimized  a  virus  purification  method  that  was  based  on  using  cell  culture  flask  and  adherent  cells.
Materials and Methods: L-929 cells were used for reovirus propagation.  After sufficient CPE, the cells were collected and pelleted.  Using  Vertrel-XF  treatment  and  ultracentrifugation  on  the  cesium  chloride  (CsCl)  gradient,  purified  reovirus  was  obtained.  It  was  subsequently  concentrated  by  filtration  using  a  100kDa  Amicon  unit.  Finally,  infectivity  and  the  number  of  purified  human  reoviruses  were  evaluated  by  plaque  assay. The  band  of  purified  human  reovirus  was  aspirated  form  the  ultracentrifugation  tube  and  then  was  dialysed  and  concentrated  by  filtration  in  Amicon  unit.  The  titer  of  purified  human  reovirus  was  determined to be  3×1012  PFUs/ml.
Results: In  present  study,  we  established  a  protocol  for  the  purification  of  human  reovirus  without  need  for  equipment  of  suspension  cell  culture. 
Conclusions: Although,  the  time-consuming  dialysis  procedure  was  removed  from  the  end  of  the  work  and  replaced  with  a  rapid  and  simple  filtration  method,  the  high  titer  of  purified  human  reovirus  was  acquired.   
Keywords: oncolytic virus, reovirus, virus purification, CsCl gradient.
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Type of Study: Original article | Subject: General
Received: 2019/12/5 | Accepted: 2020/03/31 | Published: 2020/03/31
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Hamidi-Fard M, Ataei-Pirkooh A, Aghasadeghi M, Kazemi R. Purification of Human Reovirus in Monolayer of L-929 Cells. Iran J Virol. 2019; 13 (1) :35-40
URL: http://journal.isv.org.ir/article-1-368-en.html


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