Anti-SREBP1 antibody [2A4]

• WB

Lab

Western blot - Anti-SREBP1 antibody [2A4] (AB3259)

This blot was produced using a 4-12% Bis-tris gel under the MOPS buffer system. The gel was run at 200V for 55 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was blocked for an hour using 5% milk before ab3259 and ab181602 (rabbit anti-GAPDH loading control) were incubated overnight at 4°C at a 5ug/ml concentration and 1/20000 dilution respectively. Antibody binding was detected using Goat anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed (ab216777) and Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed (ab216772) secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-SREBP1 antibody [2A4] (ab3259) at 5 µg/mL

Lane 1:

HeLa whole cell lysate at 40 µg

Lane 2:

HAP1 whole cell lysate at 40 µg

Predicted band size: 122 kDa

Observed band size: 125 kDa,65 kDa

false

• WB

CiteAb

Western blot - Anti-SREBP1 antibody [2A4] (AB3259)

Western Blotting using Anti-SREBP1 antibody [2A4], ab3259. Publication image from Han, H. J. et al., 2017, Redox Biol, 28704726. Legend direct from paper.

The regulatory role of ER stress induction by BNIP3 knock down in SREBP1 and FASN expressions. (A) UCB-hMSCs were pretreated with fatostatin (10 µM) for 30 min prior to hypoxia treatment for 24 h. The mRNA expression of FASN was analyzed by qPCR. n = 6. (B) UCB-hMSCs were transfected with BNIP3 or NT siRNAs, incubated with hypoxia for 24 h. The mRNA expression of SREBF1 was measured by qPCR. n = 6. (C) The protein expressions of mature SREBP1, FASN, BNIP3 and β-actin were detected by western blot. n = 4. (D) Cells were transfected with NT or BNIP3 siRNA for 24 h prior to hypoxia incubation for 48 h. Mature SREBP1, lamin A/C and β-tubulin expressions in non-nuclear and nuclear fractionized samples were detected by western blot. n = 3. (E) BNIP3 or NT siRNAs-transfected UCB-hMSCs were pretreated with NAC (500 µM) prior to hypoxia treatment for 48 h. Mature SREBP1, FASN, BNIP3, and β-actin were detected by western blot. n = 3. (F, G) UCB-hMSCs were transfected with BNIP3 or NT siRNAs, incubated with hypoxia for 48 h. CHOP, p-eIF2α (Ser51), eIF2α, p-mTOR (Ser2448), mTOR, p-S6K1 (Thr389), S6K1, p-S6 (Ser240/244) and β-actin expressions were shown. n = 4. (H) BNIP3 or NT siRNAs-transfected UCB-hMSCs were pretreated with NAC (500 µM) for 1 h prior to hypoxia treatment for 48 h. The expressions of CHOP, p-eIF2α (Ser51), eIF2α and β-actin were detected by western blot. n = 4. (I) Cells were transfected with BNIP3 or NT siRNAs, pretreated with PBA (100 µM) for 1 h prior to hypoxia treatment for 48 h. Mature SREBP1, FASN and β-actin proteins expressions were shown. Western blot data were normalized by β-actin, and qPCR data were normalized by ACTB mRNA expression level. Lamin A/C and β-tubulin were used as nuclear and non-nuclear protein controls, respectively. The quantitative data are presented as a mean ± S.E.M. All blot images are representative. *p < 0.05 versus control, #p < 0.05 versus hypoxia, @p < 0.05 versus BNIP3 siRNA-transfected UCB-hMSCs with hypoxia.

false