Ucp1 antibody

Manufactured by Abcam

Sourced in United Kingdom, United States

UCP1 antibody is a reagent used for the detection and analysis of the Uncoupling Protein 1 (UCP1) in biological samples. UCP1 is a mitochondrial inner membrane protein that plays a key role in thermogenesis and energy expenditure. This antibody can be utilized in various research applications, such as Western blotting, immunohistochemistry, and immunofluorescence, to investigate the expression and localization of UCP1 in different cell types and tissues.

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Lab products found in correlation

4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (3 mentions) Biotinylated goat anti rabbit secondary antibody, by Jackson ImmunoResearch (2 mentions) Dab substrate, by Vector Laboratories (1 mentions) Abc kit, by Vector Laboratories (1 mentions) Ecl western blotting substrate, by GE Healthcare (1 mentions) Fgf21, by Abcam (1 mentions) Pvdf membrane, by GE Healthcare (1 mentions) β actin antibody, by Merck Group (1 mentions) Chemidoc xrs system, by Bio-Rad (1 mentions) Chemidoc, by Bio-Rad (1 mentions) Alexa fluor 680, by Thermo Fisher Scientific (1 mentions) Hsl antibody, by Cell Signaling Technology (1 mentions) Mitochondrial total oxphos protein antibody set, by Abcam (1 mentions) Nitrocellulose membrane, by Bio-Rad (1 mentions) Dmlb microscope, by Leica (1 mentions) Affinipure goat anti rabbit igg h l, by Jackson ImmunoResearch (1 mentions) Dm6b microscope, by Leica (1 mentions) Image pro plus software, by Media Cybernetics (1 mentions) Signalstain dab reagent, by Cell Signaling Technology (1 mentions) Signalstain boost ihc detection reagent, by Cell Signaling Technology (1 mentions)

Close spelling variants of this product

Anti-UCP1 antibody (36 citations) UCP1 primary antibody (5 citations) Rabbit-anti-mouse UCP1 antibody (3 citations) UCP1 antibody (ab10983, (2 citations) Rabbit antibody against human UCP1 (2 citations)

23 protocols using ucp1 antibody

Adipose Tissue Immunohistochemical Analysis

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For immunohistochemical staining, inguinal adipose tissues were fixed in 4% paraformaldehyde for 24 h and then embedded in OCT. Cryostat sections of the inguinal adipose tissues were stained with DAPI (Eugene, Oregon, USA). The infected cells expressing GFP were visualized by immunofluorescence microscopy.
For UCP1 immunohistochemistry, 4% paraformaldehyde was used to fix inguinal adipose tissues for 24 h and then issues were embedded in paraffin. Slides (5 μm in thickness) were deparaffinized and incubated with rabbit polyclonal UCP1 antibody (1:500; Abcam, Cambridge, MA, USA) overnight at 4°C. Signals were amplified using 1:500 goat anti-rabbit immunoglobulin G horseradish peroxidase-linked antibody (Vector Laboratories, Burlingame, CA, USA) with the ABC kit (Vector Laboratories, Burlingame, CA, USA) and DAB substrate (Vector Laboratories, Burlingame, CA, USA). For H&E staining, slides were stained with eosin and hematoxylin (Nanjing Jiancheng Bioengineering Institute, Nanjing, China).

Zheng S., Guo S., Sun G., Shi Y., Wei Z., Tang Y., He F., Shi C., Dai P., Chong H., Samuelson I., Zen K., Zhang C.Y., Zhang Y., Li J, & Jiang X. (2019). Gain of Metabolic Benefit with Ablation of miR-149-3p from Subcutaneous Adipose Tissue in Diet-Induced Obese Mice. Molecular Therapy. Nucleic Acids, 18, 194-203.

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UCP1 and Glucose Transporter Proteins in Adipose Tissue

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All animals (control animals and cold-acclimated animals) were sacrificed at day 30 of the experiment. iBAT and visceral WAT were dissected and snap-frozen in liquid nitrogen. Western blot has been performed as stated previously [36 (link)]. In short, adipose tissue samples were incubated with RIPA buffer, frozen and after thawing lysates were passed through a 25-G needle. Lysates were seperated using SDS-PAGE prior to electrophoretic transfer onto nitrocellulose membranes. The UCP1 antibody was from Abcam (Cambridge, UK). CD36 and GLUT4 antibodies were purchased from Santa Cruz (Dallas, TX).

Paulus A., van Ewijk P.A., Nascimento E.B., De Saint-Hubert M., Hendrikx G., Vogg A., Pooters I., Schnijderberg M., Vanderlocht J., Bos G., Brans B., Schrauwen-Hinderling V.B., Mottaghy F.M, & Bauwens M. (2019). Characterization of BAT activity in rats using invasive and non-invasive techniques. PLoS ONE, 14(5), e0215852.

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Western Blot Analysis of Protein Targets

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Cells or tissues were lysed in RIPA buffer (Tris–HCl 50 mM, pH 7.4, NaCl 150 mM, sodium deoxycholate 0.25%, NP-40 1%, EDTA 1 mM, PMSF 1 mM, Aprotinin 1 mg/ml, leupeptin 1 mg/ml, pepstain 1 mg/ml) and a total of 20 ug of protein was separated by SDS-PAGE electrophoresis and transferred to PVDF membranes (Amersham International, GE Healthcare). Membranes were incubated with blocking solution (5% milk powder in tris-buffered saline–Tween 20 (TBST) ) for 1 h, then with primary antibody (in blocking solution) overnight at 4°C. After several washes in TBST, membranes were incubated with horseradish peroxidase (HRP)-conjugated secondary antibodies for 1 h at room temperature (RT) in blocking solution. Membranes were incubated with ECL western-blotting substrate (Amersham International, GE Healthcare) and imaged by in a ChemiDOC XRS system or ChemiDOC (Bio-Rad Laboratories).
The following antibodies were used in this study: anti-HA-tag antibody (MBL, M180–3) for CjCas9, β-tubulin mouse antibody (Proteintech, 66240-1-Ig), β-actin antibody (Sigma-Aldrich, A3854), UCP1 antibody (Abcam, ab10983), FGF21 (Abcam, ab171941).

Zhang X., Lv S., Luo Z., Hu Y., Peng X., Lv J., Zhao S., Feng J., Huang G., Wan Q.L., Liu J., Huang H., Luan B., Wang D., Zhao X., Lin Y., Zhou Q., Zhang Z.N, & Rong Z. (2021). MiniCAFE, a CRISPR/Cas9-based compact and potent transcriptional activator, elicits gene expression in vivo. Nucleic Acids Research, 49(7), 4171-4185.

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Adipose Tissue Analysis with UCP1 IHC

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Following the collection of the adipose tissues and storing in 10% NBF, they were embedded in paraffin and sectioned. H&E staining and UCP1 IHC were performed by the Advanced Molecular Pathology Laboratory (AMPL) at the Institute of Molecular and Cell Biology (IMCB), A*STAR, according to standard operating procedures. 1:100 dilution of UCP1 antibody (Abcam) was used for UCP1 IHC.

Dinish U.S., Wong C.L., Sriram S., Ong W.K., Balasundaram G., Sugii S, & Olivo M. (2017). Diffuse Optical Spectroscopy and Imaging to Detect and Quantify Adipose Tissue Browning. Scientific Reports, 7, 41357.

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Protein Expression Analysis via Western Blot

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Western blot was conducted to assess the protein of interest [23 (link)]. Snap-frozen samples were homogenized in a modified radioimmunoprecipitation assay buffer and followed by centrifugation to extract supernatants. An equal amount of protein was loaded on SDS-PAGE, which was transferred to nitrocellulose membranes (Bio-Rad, Hercules, CA, USA). The proteins were then immunoblotted with various primary and secondary antibodies, and visualized on a Li-COR Imager System (Li-COR Biosciences, Lincoln, NE, USA). The protein antibodies are listed as follows: UCP1 antibody (1:500, abcam, ab23841), Mitochondrial total OXPHOS protein antibody set (Abcam, ab110413), phospho-HSL antibody (pHSL) (1:1000, Cell Signaling 4126), HSL antibody (1:1000, Cell Signaling 4107), α-Tubulin antibody (1:1000, Advanced BioChemicals, ABCENT4777, Lawrenceville, GA, USA) and the secondary antibody Alexa Fluor 680 (ThermoFisher Scientific).

Li F., Wang S., Cui X., Jing J., Yu L., Xue B, & Shi H. (2022). Adipocyte Utx Deficiency Promotes High-Fat Diet-Induced Metabolic Dysfunction in Mice. Cells, 11(2), 181.

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Immunohistochemical Analysis of UCP-1 in Brown Adipose Tissue

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After removal of Araldite with 1% sodium hydroxide in absolute ethanol (30 min, 37ºC), semi-thin BAT sections (2 µm), were rehydrated with decreasing concentrations of ethanol, subsequently incubated in citrate buffer for 3 min at 600W to retrieve antigens and washed in phosphate buffered saline (PBS, pH 7.4). After blocking endogenous peroxidase by incubating with 3% hydrogen peroxide in methanol and three sequential washings in PBS, the sections were incubated with rabbit polyclonal UCP-1 antibody (Abcam, Cambridge, UK) overnight at 4ºC. Immunodetection was performed using the Dako LSAB Universal Kit (Dako Scientific, Glostrup, Denmark). The sections which were washed three times were incubated with 0.012% hydrogen peroxide and 0.05% diaminobenzidine (Sigma-Aldrich, Seeize, Germany) in PBS for 10 min in the dark. Finally, after rinsing in distilled water, the sections were counterstained with hematoxylin, mounted and examined with a Leica DMLB microscope (Leica Microsystems).

Golic I., Velickovic K., Markelic M., Stancic A., Jankovic A., Vucetic M., Otasevic V., Buzadzic B., Korac B, & Korac A. (2014). Calcium-Induced Alteration of Mitochondrial Morphology and Mitochondrial-Endoplasmic Reticulum Contacts in Rat Brown Adipocytes. European Journal of Histochemistry : EJH, 58(3), 2377.

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Immunohistochemical Analysis of UCP1

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Tissues were dissected and fixed in 4% neutral buffered formalin overnight and then placed in 70% ethanol for long-term storage. Paraffin processing, embedding, sectioning, H&E staining, and IHC were performed at Shanghai ZuoCheng Bio Company (Shanghai, China) according to standard protocols. The following antibodies and concentrations were used in the IHC assay: UCP1 antibody (1:50, Abcam) and peroxidase AffiniPure Goat Anti-Rabbit IgG (H + L) (1:200, Jackson ImmunoResearch). All slides were observed using a Leica DM6 B microscope at the indicated magnification, and images were captured with a sCMOS camera under the same parameter settings. Quantification of IHC images was performed using Image-Pro Plus software (Media Cybernetics Inc., Silver Springs, MD, USA).

Wu L., Xia M., Duan Y., Zhang L., Jiang H., Hu X., Yan H., Zhang Y., Gu Y., Shi H., Li J., Gao X, & Li J. (2019). Berberine promotes the recruitment and activation of brown adipose tissue in mice and humans. Cell Death & Disease, 10(6), 468.

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Immunohistochemistry of UCP1 in Adipose Tissue

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Adipose tissues were fixed in formalin for 24 h and embedded in paraffin. Tissue sections were deparaffinised, subjected to antigen unmasking (boiling in 10 mM sodium citrate pH 6 in a pressure cooker for 30 min), blocked with 5% bovine serum albumin (BSA) and incubated with UCP1 antibody (Abcam, cat. no. 10983) diluted 1:1000 for 16 h at 4 °C. Sections were then incubated with SignalStain^© Boost IHC Detection Reagent (Cell Signaling Technology, cat. no. 8114P) and immune complexes were visualised with SignalStain^© DAB reagent (Cell Signaling Technology, cat. no. 8059).

Araiz C., Yan A., Bettedi L., Samuelson I., Virtue S., McGavigan A.K., Dani C., Vidal-Puig A, & Foukas L.C. (2019). Enhanced β-adrenergic signalling underlies an age-dependent beneficial metabolic effect of PI3K p110α inactivation in adipose tissue. Nature Communications, 10, 1546.

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Immunohistochemical Analysis of UCP1 Expression

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The tissues were fixed in 10% neutral buffered formalin for 24 h and then embedded in paraffin. After deparaffinization, thin sections (4 μm) were processed for immunohistochemical staining. The slides were blocked with 1% bovine serum albumin for 1 h and were then incubated with the UCP1 antibody (1:100, Cat#: ab10983, Abcam, Cambridge, MA) at 4°C overnight. After washing off the primary antibody, sections were incubated for 1 h at room temperature with goat anti‐rabbit‐IgG‐ HRP (1:200, Cat#: sc‐2004, Santa Cruz Biotechnology, Santa Cruz, CA). The secondary antibody was washed off, followed by shaking off excess fluid. Freshly DAB Chromogenic substrate reagent (Cat#: AR1022, Boster Biological Technology, Pleasanton, CA) was added to cover the tissue section and incubated for 5 min. After rinsing with distilled water, the slide was placed in a bath of Mayer's hematoxylin for 1 min. The slide was rinsed under running tap water for 5 min and then mounted using glycerol gelatin. The images were taken with the microscope.

Yang K.T., Wang F., Lu X., Peng K., Yang T, & David Symons J. (2017). The soluble (Pro) renin receptor does not influence lithium‐induced diabetes insipidus but does provoke beiging of white adipose tissue in mice. Physiological Reports, 5(21), e13410.

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Immunohistochemical Staining of UCP1

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BAT was fixed with formalin 10%, included in paraffin, cut into 5‐μm sections, and sequentially stained with anti‐uncoupling protein 1 (UCP1) antibody (1/500, Abcam no. AB10983), biotinylated goat anti‐rabbit secondary antibody (1/500, Jackson ImmunoResearch Laboratories no. 111–066‐003), streptavidin‐conjugated avidin‐biotin complex (Vector Laboratories, no. PK‐6100), and the substrate 3,3′‐diaminobenzidene conjugated to horseradish peroxidase (Vector Laboratories, no. SK‐4100). Sections were then briefly counterstained with Nuclear Fast Red (Sigma‐Aldrich, no. N‐3020).

Crespo M., Nikolic I., Mora A., Rodríguez E., Leiva‐Vega L., Pintor‐Chocano A., Horrillo D., Hernández‐Cosido L., Torres J.L., Novoa E., Nogueiras R., Medina‐Gómez G., Marcos M., Leiva M, & Sabio G. (2023). Myeloid p38 activation maintains macrophage–liver crosstalk and BAT thermogenesis through IL‐12–FGF21 axis. Hepatology (Baltimore, Md.), 77(3), 874-887.

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