Anti gapdh

The following antibodies were used according to the protocols supplied by the manufacturers: anti-pERK1/2 (#9101, Cell Signaling Technology, Danvers, MA, USA), anti-ERK1/2 (#4695, Cell Signaling Technology), anti-GAPDH (ENM0040, Elabscience Biotechnology Inc., Houston, TX, USA), anti-pEGFR (12A3) (sc-57542, Santa Cruz Biotechnology Inc., Dallas, TX, USA), anti-EGFR (C-2) (sc-377229, Santa Cruz Biotechnology Inc.), anti-pAKT (#9271, Cell Signaling Technology), anti-AKT (#9272, Cell Signaling Technology), anti-Cyclin D1 (sc-753, Santa Cruz Biotechnology Inc.), anti-p21 (sc-397, Santa Cruz Biotechnology Inc.), and anti-beta Tubulin (Sigma), anti-alpha actin (Sigma). All the secondary antibodies (HRP-conjugated anti-rabbit and anti-mouse) were purchased from Santa Cruz Biotechnology Inc.
Drug formulations: gefitinib (ZD1839) and MG132 (S2619) were purchased from Selleckchem; AZD9291 was obtained from AstraZeneca; BAPTA_AM (HB0981) was from HelloBio.
Recombinant Human EGF (AF-100-15) was from PeproTech (London, UK).

The total protein was extracted using the kit (Beyotime Biotechnology, China), which included five samples of OSCC tissue paired with adjacent normal tissues. SDS-containing polyacrylamide gels were used to separate protein samples for transfer to polyvinylidene fluoride membranes (Millipore, USA). The membrane was treated with the primary antibody for an overnight period at 4 °C after being blocked with 5 percent BSA at room temperature. The primary antibodies were anti-p62 (1:1000; Proteintech, cat.no. 18420-1-AP), anti-LC3 (1:1000; Proteintech, cat.no. 14600-1-AP), anti-ATG5 (1:1000; Proteintech, cat.no. 10181-2-AP), anti-Bim (1:1000; Cell Signaling Technology, cat.no. #2933), anti-BCL-2 (1:1000; Proteintech, cat.no. 12789-1-AP), anti-Bax (1:1000; Cell Signaling Technology, cat.no. #14796), anti-GAPDH (1:2000; Elabscience, cat. no. E-AB-20072), and anti-Beclin-1 (1:1000; Proteintech, cat.no. 11306-1-AP). After three washes with TBST, the membrane was incubated for 1 h at room temperature with goat anti-rabbit IgG (1:10,000; Proteintech, cat.no. 10285-1-AP).