Document Detail

Centrifugal enhancement of retroviral mediated gene transfer.
MedLine Citation:
PMID:  8530565     Owner:  NLM     Status:  MEDLINE    
Centrifugation has been used for many years to enhance infection of cultured cells with a variety of different types of viruses, but it has only recently been demonstrated to be effective for retroviruses (Ho et al. (1993) J. Leukocyte Biol. 53, 208-212; Kotani et al. (1994) Hum. Gene Ther. 5, 19-28). Centrifugation was investigated as a means of increasing the transduction of a retroviral vector for gene transfer into cells with the potential for transplantation and engraftment in human patients suffering from genetic disease, i.e., gene therapy. It was found that centrifugation significantly increased the rate of transduction into adherent murine fibroblasts and into non-adherent human hematopoietic cells, including primary CD34+ enriched cells. The latter samples include cells capable of reconstitution of hematopoiesis in myeloablated patients. As a step toward optimization of this method, it was shown that effective transduction is: (1) achieved at room temperature; (2) directly related to time of centrifugation and to relative centrifugal force up to 10,000 g; (3) independent of volume of supernatant for volumes > or = 0.5 ml using non-adherent cell targets in test tubes, but dependent upon volume for coverage of adherent cell targets in flat bottom plates; and (4) inversely related to cell numbers per tube using non-adherent cells. The results support the proposal that centrifugation increases the reversible binding of virus to the cells, and together with results reported by Hodgkin et al. (Hodgkin et al. (1988) J. Virol. Methods 22, 215-230), these data support a model in which the centrifugal field counteracts forces of diffusion which lead to dissociation during the reversible phase of binding.
A B Bahnson; J T Dunigan; B E Baysal; T Mohney; R W Atchison; M T Nimgaonkar; E D Ball; J A Barranger
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Publication Detail:
Type:  Journal Article; Research Support, U.S. Gov't, P.H.S.    
Journal Detail:
Title:  Journal of virological methods     Volume:  54     ISSN:  0166-0934     ISO Abbreviation:  J. Virol. Methods     Publication Date:  1995 Aug 
Date Detail:
Created Date:  1996-01-30     Completed Date:  1996-01-30     Revised Date:  2007-11-14    
Medline Journal Info:
Nlm Unique ID:  8005839     Medline TA:  J Virol Methods     Country:  NETHERLANDS    
Other Details:
Languages:  eng     Pagination:  131-43     Citation Subset:  IM    
Department of Human Genetics, Graduate School of Public Health, University of Pittsburgh, PA 15261, USA.
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MeSH Terms
3T3 Cells
Antigens, CD34
Cell Line
Fibroblasts / cytology
Gene Transfer Techniques*
Leukemia, Erythroblastic, Acute
Retroviridae / genetics*
Tumor Cells, Cultured
Grant Support
Reg. No./Substance:
0/Antigens, CD34

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