Changes in the molecular properties of the heat-exposed cells were investigated using ICC, western blotting, and qRT-PCR. Five different primary antibodies (DAKO, Denmark),
anti-E-cadherin (1 : 300), anti-CK8/18 (1 : 200),
anti-vimentin (1 : 1000),
anti-desmin (1 : 150), and anti-p53 (1 : 300), were employed for immunocytochemistry (ICC). The antibodies were applied to quantify the changes in EMT-related properties or demonstrate the cell death mechanism.
The changes in protein expression were further determined by western blotting. The expression of heat shock protein 70 (
Hsp70, 1 : 1000, Merck Millipore, Germany),
Hsp90 (1 : 500, Santa Cruz, CA, USA),
E-cadherin (41 : 1000, Merck Millipore), and
vimentin (1 : 1000, Merck Millipore) were quantified by comparing to that of
β-actin (1 : 1000, Merck Millipore).
EMT-related genes were quantified using qRT-PCR. Primer sequences, including HSPA1A (Hs00359163_s1), CDH1 (Hs01023894_m1), VIM (Hs00958111_m1), TWIST1 (Hs01675818_s1), and GADPH (Hs02758991_g1), were purchased from Thermo Fisher Scientific. See ESI
† for additional details.
Lee T.H., Bu J., Kim B.H., Poellmann M.J., Hong S, & Hyun S.H. (2018). Sub-lethal hyperthermia promotes epithelial-to-mesenchymal-like transition of breast cancer cells: implication of the synergy between hyperthermia and chemotherapy. RSC Advances, 9(1), 52-57.