The curcumin was purchased from Shanghai Yien Chemical Technology (Shanghai, China). Rabbit polyclonal antibodies specific to CD36 (ab252922), HMGCS2 (ab137043), and ACSL1 (ab177958) were the production of Abcam (UK). CYP7A1 (DF2612), FADS2 (DF15514), and ACAA1 (DF12345) were purchased from Affinity (USA). The GAPDH (T0004) was the production of Zen Bioscience (Chengdu, China).
Ab177958
Manufactured by Abcam
Sourced in United States, United Kingdom
Ab177958 is a lab equipment product. It is a device used for research purposes in laboratory settings. The core function of this product is to facilitate specific tasks or measurements required in a laboratory environment.
Automatically generated - may contain errors
Lab products found in correlation
Pierce bca protein assay kit, by Thermo Fisher Scientific (2 mentions)
Ab252922, by Abcam (1 mentions)
Ab137043, by Abcam (1 mentions)
Ab32081, by Abcam (1 mentions)
Ab138492, by Abcam (1 mentions)
Anti erk2, by Abcam (1 mentions)
Ecl reagent, by Thermo Fisher Scientific (1 mentions)
Ab125066, by Abcam (1 mentions)
Af5186, by Affinity Biosciences (1 mentions)
Lamin b1 antibody, by Proteintech (1 mentions)
Ab23875, by Abcam (1 mentions)
Ab125247, by Abcam (1 mentions)
Ab19898, by Abcam (1 mentions)
Ab32047, by Abcam (1 mentions)
Ab46020, by Abcam (1 mentions)
Ab181557, by Abcam (1 mentions)
Ab54481, by Abcam (1 mentions)
Ab97959, by Abcam (1 mentions)
Transfection reagent, by Promega (1 mentions)
Ab39012, by Abcam (1 mentions)
9 protocols using ab177958
1
Curcumin Biomarker Antibody Validation
2
Validating Differential Protein Expression
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Western blot analysis was performed to validate the variation tendencies of DEPs identified by our proteomics approach. The protein bands were visualized by using an enhanced chemiluminescence ECL reagent (Thermo Fisher Scientific). Primary antibodies and their dilution ratio were followed, Anti-COL1A1 (1:10000, ab138492), Anti-ACSL1 (1:10000, ab177958), Anti-ERK2 (1:10000, ab32081) (Abcam, USA); HSD17β4 antibody (1:2000, 15116-1-AP), FABP4 antibody (1:1000, 12802-1-AP), PKA C-alpha antibody (1:1000, 55388-1-A), ATGL antibody (1:2000, 55190-1-AP) (Proteintech, USA), and β-actin (polyclonal antibody, 1:1000, Shenggong, China).
Immunohistochemistry staining was performed to map the distribution of FABP4 protein in the TAT according to a previously described protocol [35 (link)]. The tail adipocyte sections (thickness, 12 mm) were obtained in Tibetan sheep (TTS) and Hu sheep (FTS) and incubated overnight at 4°C with the FABP4 antibody, which was diluted 1:1000 in PBS/1% BSA, followed by incubation for 1 h at room temperature with fluorochrome-conjugated secondary antibody anti-rabbit IgG A0208 (Biyuntian, China). We performed immunofluorescence to confirm this correlation, followed by incubating in the corresponding secondary antibodies. The AlexaFluor-488 conjugated mouse IgG (Bioss, China) was incubated for 1 h at room temperature and then diluted 1:200 in PBS/1% BSA.
Immunohistochemistry staining was performed to map the distribution of FABP4 protein in the TAT according to a previously described protocol [35 (link)]. The tail adipocyte sections (thickness, 12 mm) were obtained in Tibetan sheep (TTS) and Hu sheep (FTS) and incubated overnight at 4°C with the FABP4 antibody, which was diluted 1:1000 in PBS/1% BSA, followed by incubation for 1 h at room temperature with fluorochrome-conjugated secondary antibody anti-rabbit IgG A0208 (Biyuntian, China). We performed immunofluorescence to confirm this correlation, followed by incubating in the corresponding secondary antibodies. The AlexaFluor-488 conjugated mouse IgG (Bioss, China) was incubated for 1 h at room temperature and then diluted 1:200 in PBS/1% BSA.
3
Ferroptosis Pathway Regulation Assay
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Sigma‐Aldrich (MO, USA) supplied Fer‐1 and erastin, while Proteintech Group, Inc. provided the primary antibodies: anti‐YAP (20536‐1‐AP,1:1000), anti‐RUNX2 (AF5186,1:1000; Affinity Biosciences), anti‐TEAD2 (21159‐1‐AP,1:1000; Proteintech Group, Inc.), Lamin B1 antibody (12987‐1‐AP,1:1000; Proteintech Group, Inc.), anti‐β‐catenin (20536‐1‐AP,1:1000; Proteintech Group, Inc.), anti‐GPX4 (ab125066,1:1000; Abcam) and anti‐ACSL4 (ab177958,1:1000; Abcam). Cell Signaling Technology (MA, USA) supplied all the secondary antibodies.
4
Stem Cell Protein Expression Analysis
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According to previous description, western blot was conducted44 (link). Following primary antibodies were acquired from Abcam (Cambridge, USA): SOX2 (ab97959), Oct4 (ab181557), LIN28 (ab46020), CD133 (ab19898), CPT1 (ab189182), ACS (ab177958), p-AMPK (ab23875), AMPK (ab32047), PGC1α (ab54481), GAPDH (ab125247).
5
Liver Cell Culture and ACSL1 Antibody
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Human liver cells and 1640 medium were purchased from Zhongqiao Xinzhou Biotechnology Co. Ltd (Shanghai, China). Australian fetal bovine serum and dual antibody were purchased from Invitrogen, Carlsbad, CA, USA and transfection reagents were purchased from Promega (Beijing) Biotech Co., Ltd. ACSL1 monoclonal antibody (1: 1000, ab177958) was purchased from Abcam, Cambridge, MA, USA.
6
Protein Profiling via RIPA Lysis and Western Blot
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Radioimmunoprecipitation assay (RIPA) solution encompassing protease inhibitors (Solarbio) was used to lysate total cell proteins. The proteins were then transferred to polyvinylidene difluoride (PVDF) membranes (Roche, Basel, Switzerland) after being separated on an SDS–polyacrylamide gel. At room temperature, the membranes were blocked with 5% fat-free dry milk before being cultivated with primary antibodies with diluted 2000 times. The membranes were then treated with the secondary antibody for 2 h at room temperature. ECL-enhanced chemiluminescence reagents (Amersham, Little Chalfont, UK) were used to visualize protein bands. Primary antibodies used in this investigation included anti-PDK4 (ab110336; Abcam, Cambridge, UK), anti-MMP-3 (ab52915; Abcam), anti-MMP-13 (ab39012; Abcam), anti-ADAMTS-4 (ab314856; Abcam), anti-PPARA (ab227074; Abcam), anti-PPARD (ab178866; Abcam), anti-ACSL1 (ab177958; Abcam), and anti-GAPDH (ab52915; Abcam).
7
NLRP3 Inflammasome Immunohistochemistry
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The paraffined sections were dewaxed, dehydrated with gradient ethanol, and washed with PBS (three times/3 min). Then, the sections were cultured with 3 ml/L methanol-H2O2 for 15 min to block endogenous peroxidase, followed by PBS washing (three times/3 min). Next, the sections were detached with 1 g/L trypsin at 37°C for 30 min to expose intracellular antigens, followed by PBS washing (three times/3 min). After that, the sections were blocked with skim milk powder and cultured with the primary antibodies: anti-NLRP3 (ab214185, 1/100, Abcam Inc., Cambridge, MA, USA), anti-ACS (ab177958, 1/200, Abcam), anti-caspase-1 (ab74279, 1/250, Abcam), and anti-IgG (ab172730, 1/500, Abcam) at 4 °C overnight. Following PBS washing (three times/3 min), the sections were cultured with the secondary antibody immunoglobulin G (IgG) (ab6721, 1/1000, Abcam) at 37°C for 1 h. Thereafter, the sections were developed with 2,4-diaminobutyric acid (DAB) and counterstained with hematoxylin for 3–5 min. Afterwards, the sections were dehydrated, dried and sealed.
8
Comprehensive Aortic Root Histological Analysis
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Mice treated with the indicated treatments were euthanized with pentobarbital sodium (100 mg/kg). Tissues were fixed in 4% PFA, mounted in paraffin blocks, and sliced at 8 μm. Frozen aortic roots were dissected for serial 8 μm cryo-sectioning that covered 500 μm of the root. Aortic root sections were stained with hematoxylin and eosin (H&E), Oil Red O (Sigma), Masson's trichrome staining (Solarbio, China), and anti-ACSL1 (Abcam, ab177958) antibodies. Furthermore, quantitative analysis of atherosclerotic plaques was performed using Image-Pro Plus 6.0. For immunofluorescence analysis, mice were sacrificed, and tissues of interest were dissected. Tissues were fixed in 4% PFA, mounted in paraffin blocks, and sliced into 8 μm sections. CD68+iNOS+ and CD68 + ARG1+ cells in aortic roots were double stained with corresponding antibodies: CD68 (Abcam, ab53444), iNOS (Abcam, ab178945), and ARG1 (Abcam, ab91279), followed by incubation with specific fluorescence-labeled secondary antibodies for 2 h. Cell nuclei were counter-stained with Hoechst33342 for 15 min at room temperature in the dark. Images were captured using a laser scanning confocal microscope (Nikon A1R, Tokyo, Japan).
9
Protein Expression Analysis by Western Blot
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Tissues and cells were harvested in 1 × RIPA buffer containing a protease inhibitor (Beijing, China). Samples were shaken on a rotator for 30 min and subsequently centrifuged at 12,000 rpm for 20 min at 4 °C. Protein concentration was determined using the Pierce BCA Protein Assay Kit (Thermo, USA). Protein samples were separated by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and transferred to a nitrocellulose membrane in an ice bath. Membrane was blocked with 5% bovine serum albumin for 1 h and incubated overnight with primary antibodies at 4 °C. The antibodies used were rabbit anti-ACSL1 (Abcam, ab177958) and rabbit anti-GAPDH (Abcam, ab181602). Membranes were then incubated with the corresponding secondary antibodies for 1 h at room temperature. The bands were visualized using ECL Prime western blotting Detection Reagent (GE Healthcare, Buckinghamshire, UK).
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