Baculoviruses
carrying either
the reference or variant SLCO1B1 and sodium butyrate
(5 mM final concentration) were added to the HEK293 cells after the
cells were cultured for 24 h in T175 flasks. After 48 h of culturing,
the cells were pelleted by centrifugation (3000g,
15 min) and broken down using a Dounce tissue homogenizer and resuspended
in Tris-sucrose (TS) buffer (10 mM Tris-HEPES, 250 mM sucrose, pH
7.4) while being kept on ice. The cell homogenate was centrifuged
for 30 min (3220 g, 4 °C), separating the larger cellular organelles
and the nucleus in the pellet. The resulting supernatant was subsequently
separated into new tubes and centrifuged (21,000g, 4 °C, 99 min) again, resulting in a pellet containing the
crude cell membrane. The protein sample was suspended in TS buffer,
and the total protein concentration was quantified as previously described.
carrying either
the reference or variant SLCO1B1 and sodium butyrate
(5 mM final concentration) were added to the HEK293 cells after the
cells were cultured for 24 h in T175 flasks. After 48 h of culturing,
the cells were pelleted by centrifugation (3000g,
15 min) and broken down using a Dounce tissue homogenizer and resuspended
in Tris-sucrose (TS) buffer (10 mM Tris-HEPES, 250 mM sucrose, pH
7.4) while being kept on ice. The cell homogenate was centrifuged
for 30 min (3220 g, 4 °C), separating the larger cellular organelles
and the nucleus in the pellet. The resulting supernatant was subsequently
separated into new tubes and centrifuged (21,000g, 4 °C, 99 min) again, resulting in a pellet containing the
crude cell membrane. The protein sample was suspended in TS buffer,
and the total protein concentration was quantified as previously described.
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