Several proteins mediate the biosynthesis of cholesterol. The first specific step in the cholesterol biosynthetic pathway is the conversion of transfarnesyl-diphosphate to Squalene, which is catalyzed by the endoplasmic reticulum membrane-associated enzyme Squalene synthetase, also designated Squalene synthase and Farnesyl-diphosphate farnesyltransferase. Squalene synthetase is located at a branch point in the mevalonate pathway and is also involved in isoprenoid biosynthesis. Squalene epoxidase, also designated Squalene monooxygenase, is a multi-pass microsomal membrane-associated enzyme that catalyzes the first oxygenation step in sterol biosynthesis and most likely functions as one of the rate-limiting enzymes in this pathway. Squalene epoxidase may form a complex with Squalene synthetase.
Background References
1. Dong X. et. al. Bavachinin inhibits cholesterol synthesis enzyme FDFT1 expression via AKT/mTOR/SREBP-2 pathway. Int Immunopharmacol. 2020 Nov
2. Lu J. et. al. 20(S)-Rg3 upregulates FDFT1 via reducing miR-4425 to inhibit ovarian cancer progression. Arch Biochem Biophys. 2020 Oct
Western blot analysis of FDFT1 on different lysates with Rabbit anti-FDFT1 antibody (HA720048) at 1/1,000 dilution.
Lane 1: HCT 116-si NT cell lysate Lane 2: HCT 116-si FDFT1 cell lysate
Lysates/proteins at 10 µg/Lane.
Predicted band size: 48 kDa Observed band size: 45 kDa
Exposure time: 10 seconds; ECL: K1801;
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 (HA720048) at 1/1,000 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 FDFT1 on HCT116 cell lysate with Rabbit anti-FDFT1 antibody (HA720048) at 1/5,000 dilution.
Lysates/proteins at 10 µg/Lane. Exposure time: 20 seconds; ECL: K1801
Blocking: 5% NFDM/TBST, 1 hour at room temperature Primary antibody: HA720048, 1/5,000 in primary antibody dilution buffer (K1803), overnight at 4 ℃ Secondary antibody: Goat anti-Rabbit IgG-HRP (HA1001), 1/50,000 in 5% NFDM/TBST, 1 hour at room temperature
Predicted band size: 48.1 kDa Observed band size: 48 kDa
Western blot analysis of FDFT1 on rat kidney tissue lysate with Rabbit anti-FDFT1 antibody (HA720048) at 1/5,000 dilution.
Lysates/proteins at 15 µg/Lane. Exposure time: 62 seconds; ECL: K1801
Blocking: 5% NFDM/TBST, 1 hour at room temperature Primary antibody: HA720048, 1/5,000 in primary antibody dilution buffer (K1803), overnight at 4 ℃ Secondary antibody: Goat anti-Rabbit IgG-HRP (HA1001), 1/50,000 in 5% NFDM/TBST, 1 hour at room temperature
Predicted band size: 48 kDa Observed band size: 45 kDa
Western blot analysis of FDFT1 on mouse kidney tissue lysate with Rabbit anti-FDFT1 antibody (HA720048) at 1/5,000 dilution.
Lysates/proteins at 15 µg/Lane. Exposure time: 62 seconds; ECL: K1801
Blocking: 5% NFDM/TBST, 1 hour at room temperature Primary antibody: HA720048, 1/5,000 in primary antibody dilution buffer (K1803), overnight at 4 ℃ Secondary antibody: Goat anti-Rabbit IgG-HRP (HA1001), 1/50,000 in 5% NFDM/TBST, 1 hour at room temperature
Predicted band size: 48 kDa Observed band size: 45 kDa
Immunohistochemical analysis of paraffin-embedded rat liver tissue using anti-FDFT1 antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA720048, 1/200) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
Immunohistochemical analysis of paraffin-embedded human liver tissue using anti-FDFT1 antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA720048, 1/50) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
Immunohistochemical analysis of paraffin-embedded human liver carcinoma tissue using anti-FDFT1 antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA720048, 1/50) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
ICC staining of FDFT1 in SH-SY5Y cells (green). Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 10% negative goat serum for 15 minutes at room temperature. Cells were probed with the primary antibody (HA720048, 1/50) for 1 hour at room temperature, washed with PBS. Alexa Fluor®488 conjugate-Goat anti-Rabbit IgG was used as the secondary antibody at 1/1,000 dilution. The nuclear counter stain is DAPI (blue).
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