Lipid extraction from the conditioned media of BMMCs

HM Hidenori Moriyama
JE Jin Endo
MK Masaharu Kataoka
YS Yuta Shimanaka
NK Nozomu Kono
YS Yuki Sugiura
SG Shinichi Goto
HK Hiroki Kitakata
TH Takahiro Hiraide
NY Naohiro Yoshida
SI Sarasa Isobe
TY Tsunehisa Yamamoto
KS Kohsuke Shirakawa
AA Atsushi Anzai
YK Yoshinori Katsumata
MS Makoto Suematsu
KK Kenjiro Kosaki
KF Keiichi Fukuda
HA Hiroyuki Arai
MS Motoaki Sano
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To obtain conditioned media from BMMCs, 1 × 107 BMMCs were cultured in IL-3-containing BMMC complete medium for 2 days. Neutral lipids, phospholipids and free fatty acids were extracted from the conditioned media by solid-phase extraction using Sep-Pak C18 cartridges (Waters). Briefly, the conditioned medium was added to Sep-Pak columns and neural lipids were eluted with 10 ml of hexane. Free fatty acids were subsequently eluted with 10 ml of methyl formate. Phospholipids were finally eluted with 10 ml of methanol. The free fatty acid fraction was used as experiments.

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