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Biotechnology and Applied Biochemistry (2010) 55, (1–8) (Printed in Great Britain)
A novel harvesting method for cultured cells using iron-cross-linked alginate films as culture substrates
Ikuko Machida‐Sano1, Yasushi Matsuda and Hideo Namiki
Department of Biology, School of Education, Waseda University, 2-2 Wakamatsu-cho, Shinjuku-ku, Tokyo 162-8480, Japan
Key words: cell cryopreservation, cell harvesting, cell subculture, ferric-ion-cross-linked alginate (Fe-alginate), scraping, trypsinization. Abbreviations used: E-MEM, Eagle's minimum essential medium; FBS, fetal bovine serum; Fe-alginate, ferric-ion-cross-linked alginate; SEM, scanning electron microscopy; TCP, tissue culture polystyrene; WST-8, 2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H‐tetrazolium, monosodium salt. 1To whom correspondence should be addressed (email ikuko@ruri.waseda.jp). The present study was conducted to assess the efficiency of a novel cell-harvesting method involving dissolution of the culture substrate composed of Fe-alginate (ferric-ion-cross-linked alginate). Cell harvesting is an essential step for recovery of cultured adherent cells, but conventional methods such as trypsinization or scraping cause considerable damage to the cells. We therefore devised an original method for harvesting cultured cells using Fe-alginate films as a culture substrate and then retrieving the cells by disintegration of the alginate gel. Fe-alginate was easily dissolved under physiological conditions by exchange of cross-linked ions using chelating agents such as citrate. The effects of this cell-harvesting method were investigated in comparison with trypsinization and scraping. Cells harvested by dissolution of the Fe-alginate substrate showed high viability and metabolic activity, high recovery rate on subculture, a low degree of membrane damage with superior expression of cell adhesion molecules (β1 integrin and N-cadherin), high resistance to oxidative stress, and high viability, metabolic activity and recovery rate after cryopreservation, in contrast with the cells harvested by trypsinization or scraping from tissue-culture polystyrene substrates. These results suggest that our new system involving dissolution of Fe-alginate culture substrate is effective for cell harvesting and may be advantageous for biomedical applications. Received 29 July 2009/5 November 2009; accepted 11 November 2009 Published as Immediate Publication 11 November 2009, doi:10.1042/BA20090215 © 2010 Portland Press Ltd |
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