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Spheroid formation

DM Daniel Martin
MD Maria S. Degese
LV Lynn Vitale-Cross
RI Ramiro Iglesias-Bartolome
JV Juan Luis Callejas Valera
ZW Zhiyong Wang
XF Xiaodong Feng
HY Huwate Yeerna
VV Vachan Vadmal
TM Toshiro Moroishi
RT Rick F. Thorne
MZ Moraima Zaida
BS Bradford Siegele
SC Sok C. Cheong
AM Alfredo A. Molinolo
YS Yardena Samuels
PT Pablo Tamayo
KG Kun Liang Guan
SL Scott M. Lippman
JL J. Guy Lyons
JG J. Silvio Gutkind
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Spheroid (orosphere) assays were performed in 24-well plates. Briefly, the wells were coated with 250 µL, 1.5%, agarose in DMEM. Ten thousand cells were resuspended in 0.5 mL warm DMEM, 10% FBS, and 0.1% agarose containing the appropriate treatment and layered on top. Agarose was allowed to solidify at room temperature for 15 min before adding an additional 0.5 mL of DMEM, 10% FBS containing the same treatment as above, and returning the cells to the incubator. The cells were allowed to grow for two weeks, replacing the liquid media every three days before spheroid formation was analyzed. For quantification, the wells were photographed using an automatic whole well scan mode in a Zeiss Axiovert 200 M microscope. Colony counting and diameter measurements were performed manually using the Zeiss Axiovision 4.8 software.

Copyright and License information: The Author(s) ©2018
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