Lactate dehydrogenase (LDH) assay (Koreniewski & Callewaert,
1983) was modified according to our previous paper (Yamashoji &
Matsuda, 2013). C6 cells prepared under the above conditions were
incubated with 100 lL of Medium A containing a-tomatine for 4 h.
After the incubation, 50 lL of the supernatant in each well of a
96-well plate were transferred to each well of another plate, and
were mixed with 50 lL of LDH assay reagent prepared with
LDH-Cytotoxic Test Wako (Wako Pure Chemical Industries, Ltd.,
Osaka, Japan). The absorbance at 560 nm was determined after
10 min-incubation with the above reagent. The viability was
calculated by the following equation.
Viability ð%Þ¼100½ðabsorbance of the cells incubated with
a-tomatineabsorbance of the untreated cellsÞ
=ðabsorbance of the cells incubated with Tween 20
absorbance of the untreated cellsÞ ed wit: