In situ brdu red dna fragmentation tunel assay kit

Manufactured by Abcam

Sourced in United States, United Kingdom

The In situ BrdU-Red DNA fragmentation (TUNEL) assay kit is a laboratory product used to detect and quantify apoptotic cells. The kit utilizes the terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) method to label DNA strand breaks and the incorporation of bromodeoxyuridine (BrdU) to visualize the labeled DNA. The core function of this kit is to facilitate the identification and analysis of cells undergoing programmed cell death.

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Lab products found in correlation

Fitc annexin 5 apoptosis detection kit 1, by BD (2 mentions) 4 6 diamidino 2 phenylindole (dapi), by Merck Group (2 mentions) Vectashield with dapi, by Vector Laboratories (1 mentions) Tissue tek optimal cutting temperature compound, by Sakura Finetek (1 mentions) Cleaved caspase 3, by Cell Signaling Technology (1 mentions) Cleaved parp, by Cell Signaling Technology (1 mentions) Sqstm1 p62, by Cell Signaling Technology (1 mentions) Propidium iodide, by Merck Group (1 mentions) 5 fluorouracil, by Merck Group (1 mentions) Chloroquine, by Merck Group (1 mentions) Halofuginone hydrobromide, by Merck Group (1 mentions) β actin, by Cell Signaling Technology (1 mentions) Goat anti rabbit hrp secondary antibody, by Thermo Fisher Scientific (1 mentions) Lc3 2, by Cell Signaling Technology (1 mentions) Z lehd fmk, by R&D Systems (1 mentions) Pierce r bca protein assay kit, by Merck Group (1 mentions) Caspase 8 inhibitor z ietd fmk, by R&D Systems (1 mentions) Artemisinin, by Abcam (1 mentions) Cleaved capase 9, by Cell Signaling Technology (1 mentions) Earle s balanced salt solution, by Merck Group (1 mentions)

Close spelling variants of this product

TUNEL kit (28 citations) TUNEL assay (27 citations) BrdU assay (8 citations) BrdU-Red (4 citations) BrdU-Red DNA fragmentation assay kit (3 citations) In situ BrdU-Red DNA Fragmentation Assay Kit (2 citations)

19 protocols using in situ brdu red dna fragmentation tunel assay kit

Apoptosis Detection in Brain Cryosections

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Identification of apoptotic cells in brain cryosections was performed using In Situ BrdU-Red DNA fragmentation (TUNEL) assay kit (Abcam). Sections were mounted using Vectashield with DAPI (Vector Laboratories).

Atkins A., Xu M.J., Li M., Rogers N.P., Pryzhkova M.V, & Jordan P.W. (2020). SMC5/6 is required for replication fork stability and faithful chromosome segregation during neurogenesis. eLife, 9, e61171.

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Apoptosis Detection in Xenograft Tissues

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Control and treated xenograft tumor tissues were placed in frozen tissue matrix and serial sections 5 microns thick were placed onto slides and stained for TUNEL assay. Sections were deparaffinized and apoptotic cells were detected using the in situ BrdU-Red DNA fragmentation (TUNEL) assay kit (Abcam) and counterstained with DAPI.

Dilly A.K., Honick B.D., Lee Y.J., Guo Z.S., Zeh H.J., Bartlett D.L, & Choudry H.A. (2017). Targeting G-protein coupled receptor-related signaling pathway in a murine xenograft model of appendiceal pseudomyxoma peritonei. Oncotarget, 8(63), 106888-106900.

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Histological Analysis of Testicular Apoptosis

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Testes were either fixed in Bouins fixative or cryopreserved using Tissue-Tek optimal cutting temperature compound (O.C.T.; Sakura Finetek). Fixed tissues were embedded in paraffin and serial sections of 5-μm thickness were placed onto slides and stained with hematoxylin and eosin or PAS. For the TUNEL assay, sections were deparaffinized and apoptotic cells were detected using the in situ BrdU-Red DNA fragmentation (TUNEL) assay kit (Abcam) and counterstained with 4′,6-diamidino-2-phenylindole (DAPI). Cyropreserved testes were sectioned at a 5-μm thickness in series using a Cryostat (Thermo Scientific CyroStar NX70). Cryopreserved testes were subsequently fixed (1% paraformaldehyde [PFA], 0.1% Triton X in 1× phosphate-buffered saline [PBS]), and subjected to standard immunostaining procedures. Primary antibodies and dilution used are presented in Supplemental Table S2. Secondary antibodies against human, rabbit, rat, mouse, and guinea pig immunoglobulin G (IgG) and conjugated to Alexa 350, 488, 568, or 633 (Life Technologies) were used at 1:500 dilution.

Hwang G., Verver D.E., Handel M.A., Hamer G, & Jordan P.W. (2018). Depletion of SMC5/6 sensitizes male germ cells to DNA damage. Molecular Biology of the Cell, 29(25), 3003-3016.

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Quantification of Neurodegeneration Markers

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Immunofluorescence analyses were performed on freshly frozen, OCT-embedded and sectioned slides (10 μm). 6 mice from ApoE^−/− and ApoE^−/−/Tollip^−/− mice were used for the study. TUNEL stainings were performed with in situ BrdU-Red DNA Fragmentation (TUNEL) assay Kit according as the product protocol (Abcam). The average numbers of positive staining cells per viewing field were quantified from 2-4 slides collected from each of the six mice.
For the measurement of β-amyloid, α-synuclein, 2-4 Slides from each mouse brain were fixed in 4% neutrally buffered formalin for 5 min, and stained with anti-mouse primary antibodies (1:100, anti-mouse β-amyloid, α-synuclein antibodies, as well as isotype control antibodies) followed by a biotinylated anti-Ig secondary Ab (BD Biosciences) and streptavidin-PE or FITC. DAPI was used to stain nucleus. 2-4 viewing fields from each slide were captured under fluorescent microscope. Pixel values reflecting the fluorescent intensities of each viewing field were quantitated with the NIH ImageJ software.

Chen K., Yuan R., Geng S., Zhang Y., Ran T., Kowalski E., Liu J, & Li L. (2016). Toll-interacting protein deficiency promotes neurodegeneration via impeding autophagy completion in high-fat diet-fed ApoE−/− mouse model. Brain, behavior, and immunity, 59, 200-210.

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Apoptosis Index Evaluation in Tissues

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The apoptosis index in the tissues of treated mice was evaluated using in situ BrdU-Red DNA fragmentation (TUNEL) assay kit from the Abcam following the manufacturer’s instructions. Briefly, 3–5 μm thick sections of paraffin-embedded tissue on glass slides were deparaffinized and hydrated. The slides were then immersed sequentially in 0.85% NaCl and PBS, followed by antigen retrieval by proteinase K (10 mg/mL) in Tris-HCl pH 8.0 + 50 mM EDTA. The tissue samples were then labeled with DNA labeling solution for 1 hour at 37°C following the fixation with 4% paraformaldehyde. After washing, the samples were incubated with anti-BrdU-Red antibody and analyzed by confocal microscopy using 20× magnification objective, after washing and mounting with fluoroshield solution.

Khan A., Alhumaydhi F.A., Alwashmi A.S., Allemailem K.S., Alsahli M.A., Alrumaihi F.A., Almatroudi A., Mobark M.A., Mousa A, & Khan M.A. (2020). Diallyl Sulfide-Mediated Modulation of the Fatty Acid Synthase (FASN) Leads to Cancer Cell Death in BaP-Induced Lung Carcinogenesis in Swiss Mice. Journal of Inflammation Research, 13, 1075-1087.

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Apoptosis and Autophagy Analysis

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Halofuginonehy drobromide, 5-fluorouracil, Earle's Balanced Salt Solution (EBSS), chloroquine (CQ), propidium iodide (PI) and Pierce (R) BCA Protein Assay Kit were obtained from Sigma-Aldrich (Munich, Germany). Artemisinin and In situ BrdU-red DNA fragmentation (TUNEL) assay kit were purchased from Abcam (Cambridge, UK). Antibodies against cleaved capase-8, cleaved capase-9, cleaved caspase-3, cleaved PARP, SQSTM1/p62, LC3-II and β-Actin were purchased form Cell Signaling Technology (Danvers, MA, USA). HRP-goat anti-rabbit secondary antibody was purchased from Invitrogen (Carlsbad, CA, USA). Goat antimouse IgG-HRP secondary antibody and caspase-3 inhibitor (z-DEVD-fmk) were purchased from San Cruz Biotechnology (Santa Cruz, CA, USA). Caspase-8 inhibitor (z-IETD-fmk) and caspase-9 inhibitor (z-LEHD-fmk) were purchased from R&D Systems (Wiesbaden-Nordenstedt, Germany). FITC Annexin V Apoptosis Detection Kit I was obtained from BD Bioscience (San Jose, CA, USA).

Gong R.H., Yang D.J., Kwan H.Y., Lyu A.P., Chen G.Q, & Bian Z.X. (2022). Cell death mechanisms induced by synergistic effects of halofuginone and artemisinin in colorectal cancer cells. International Journal of Medical Sciences, 19(1), 175-185.

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Halofuginone Cytotoxicity and Apoptosis Assay

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Halofuginone hydrobromide, 2′, 7′-dichlorodihydrofluorescin diacetate (DCFH-DA), Collagenase A, Dnase I, HPLC-grade acetonitrile, HPLC-grade methanol, HPLC-grade isopropanol, Methyl tertbutyl ether (MTBE), Ammonium acetate, N-Acetyl-L-cysteine, acetic acid, Dulbecco's Modified Eagle's Medium without glucose, L-glutamine, phenol red, sodium pyruvate and sodium bicarbonate (DMEM, D5030) and Pierce (R) BCA Protein Assay Kit and [NADP⁺]/[NADPH] Quantification Kit were obtained from Sigma-Aldrich (Munich, Germany). Rat EGF was purchased from Pepro Tech (Rocky Hill, USA). [U-¹³C₆]-glucose was purchased from Cambridge Isotope Laboratories (Tewksbury, USA). FITC Annexin V Apoptosis Detection Kit I was obtained from BD Bioscience (USA). Dialyzed Fetal Bovine Serum (US Origin SH30079.03) was purchased from HyClone (USA). In situ BrdU-red DNA fragmentation (TUNEL) assay kit (ab66110) was obtained from Abcam.

Chen G.Q., Tang C.F., Shi X.K., Lin C.Y., Fatima S., Pan X.H., Yang D.J., Zhang G., Lu A.P., Lin S.H, & Bian Z.X. (2015). Halofuginone inhibits colorectal cancer growth through suppression of Akt/mTORC1 signaling and glucose metabolism. Oncotarget, 6(27), 24148-24162.

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TUNEL Assay for Apoptosis Detection

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For the TUNEL assay, sections were processed using the in situ BrdU‐Red DNA fragmentation (TUNEL) assay kit (Abcam) and counterstained with DAPI. Spinal cord sections cut from PFA fixed tissue blocks (10–12 μm) were dried, immersed in 10 mM Tris‐HCI (pH = 7.4) at 37°C for 5 min, and then incubated in 20 μg/ml Proteinase K solution (2 μl Proteinase K 10 mg/ml + 998 μl 10 mM Tris‐HCl pH = 8.0 containing 50 mM Ethylenediaminetetraacetic acid [EDTA]) for 5 min, washed and incubated in PFA for 5 min. Tissues were covered with DNA labelling solution and incubated 1 h in the dark in a humidified 37°C incubator. Sections were then washed and incubated with anti‐BrdU‐Red antibody for 30 min, then incubated with 7‐AAD/RNase A Staining Buffer for optional counterstain for total DNA for 30 min. Cell nuclei were visualized with DAPI (Sigma‐Aldrich). Microscopy and image acquisition were performed as above. The total number of TUNEL‐positive cells per lumbar spinal cord section was counted in four different sections, averaged and compared between genotypes.

Papaneophytou C.P., Georgiou E., Karaiskos C., Sargiannidou I., Markoullis K., Freidin M.M., Abrams C.K, & Kleopa K.A. (2018). Regulatory role of oligodendrocyte gap junctions in inflammatory demyelination. Glia, 66(12), 2589-2603.

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Quantifying Apoptosis: In Vitro and In Vivo Assays

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For in vitro apoptosis detection, the Cell Death Detection Elisa^PLUS kit (Roche) was used to detect apoptosis following the manufacturer’s protocol. This assay determines apoptosis by measuring mono- and oligonucleosomes in the lysates of apoptotic cells. The cell lysates were placed into a streptavidin-coated microplate and incubated with a mixture of anti-histone-biotin and anti-DNA-peroxidase. The amount of peroxidase retained in the immunocomplex was photometrically determined with ABTS as the substrate. Absorbance was measured at 405 nm. For in vivo apoptosis detection, the in situ BrdU-Red DNA Fragmentation (TUNEL) Assay Kit (Abcam, Cambridge, MA, USA) was used following the manufacturer’s protocol. This assay utilizes Br-dUTP (bromolated deoxyuridine triphosphate nucleotides), which is readily incorporated into DNA strand breaks. The Br-dUTP sites are identified by a red fluorescence labeled anti-BrdU monoclonal antibody.

Zhang Q., Sun M., Zhou S, & Guo B. (2016). Class I HDAC inhibitor mocetinostat induces apoptosis by activation of miR-31 expression and suppression of E2F6. Cell Death Discovery, 2, 16036-.

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TUNEL Assay for Apoptosis Detection

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For the TUNEL assay, apoptotic cells were detected using the in situ BrdU-Red DNA fragmentation (TUNEL) assay kit (Abcam, Cambridge, UK) and counterstained with 4',6'-diamidino-2-phenylindole hydrochloride (DAPI).

Ren J., Kong W., Lu F, & Li Y. (2021). Adipose-derived stem cells (ADSCs) inhibit the expression of anti-apoptosis proteins through up-regulation of ATF4 on breast cancer cells. Annals of Translational Medicine, 9(16), 1300.

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