Western blot analysis of GRP78 BiP on different lysates with Mouse anti-GRP78 BiP antibody (HA601076) at 1/1;000 dilution. Lane 1: HepG2 cell lysate (10 µg/Lane) Lane 2: MCF-7 cell lysate (10 µg/Lane) Lane 3: RAW264.7 cell lysate (10 µg/Lane) Lane 4: F9 cell lysate (10 µg/Lane) Lane 5: NIH/3T3 cell lysate (10 µg/Lane) Lane 6: PC-12 cell lysate (10 µg/Lane) Lane 7: PMVEC cell lysate (10 µg/Lane) Lane 8: Mouse brain tissue lysate (20 µg/Lane) Predicted band size: 72 kDa Observed band size: 75 kDa Exposure time: 1 minute; 8% SDS-PAGE gel. Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA601076) at 1/1;000 dilution was used in 5% NFDM/TBST at room temperature for 2 hours. Goat Anti-Mouse IgG - HRP Secondary Antibody (HA1006) at 1:100;000 dilution was used for 1 hour at room temperature.
Western blot analysis of EIF2S1 on different lysates with Rabbit anti-EIF2S1 antibody (HA500385) at 1/500 dilution. Lane 1: Mouse colon tissue lysate; 20 µg/Lane Lane 2: Rat spleen tissue lysate; 20 µg/Lane Predicted band size: 36 kDa Observed band size: 36 kDa Exposure time: 30 seconds; 12% SDS-PAGE gel. Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA500385) at 1/500 dilution was used in 5% NFDM/TBST at room temperature for 2 hours. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1:300;000 dilution was used for 1 hour at room temperature.
Western blot analysis of Phospho-EIF2S1 (S51) on different lysates with Rabbit anti-Phospho-EIF2S1 (S51) antibody (ET1603-14) at 1/2;000 dilution. Lane 1: HeLa whole cell lysate (15 µg/Lane) Lane 2: HeLa treated with 50nM Calyculin A for 3 hours whole cell lysate (15 µg/Lane) Lane 3: NIH/3T3 whole cell lysate (15 µg/Lane) Lane 4: NIH/3T3 treated with 100nM Calyculin A for 30 minutes whole cell lysate (15 µg/Lane) Lane 5: C6 whole cell lysate (20 µg/Lane) Lane 6: C6 treated with 100nM Calyculin A for 30 minutes whole cell lysate (20 µg/Lane) Predicted band size: 36 kDa Observed band size: 36 kDa Exposure time: Lane 1-4: 2 minutes; Lane 5-6: 23 seconds； 4-20% SDS-PAGE gel. Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (ET1603-14) at 1/2;000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1:50;000 dilution was used for 1 hour at room temperature.
Western blot analysis of ATG12 on different lysates with Rabbit anti-ATG12 antibody (HA721504) at 1/1;000 dilution. Lane 1: HeLa cell lysate; 20 µg/Lane Lane 2: HCT 116 cell lysate; 20 µg/Lane Lane 3: HEK-293 cell lysate; 20 µg/Lane Lane 4: SH-SY5Y cell lysate; 20 µg/Lane Predicted band size: 15 kDa Observed band size: 55/20 kDa Exposure time: 3 minutes; 4-20% SDS-PAGE gel. Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA721504) at 1/1;000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1:100;000 dilution was used for 1 hour at room temperature.
