The cancer tissues of nude mice were fixed in 4% formalin and embedded in paraffin. Then, the cell apoptosis was examined using a TUNEL apoptosis detection kit (KeyGen, Nanjing, China). The apoptotic cells were captured and counted in five random fields (magnification, ×200) for each group. The apoptotic index of the cancer cells = apoptotic cells/total cells × 100%.
Tunel apoptosis detection kit
Manufactured by Keygen Biotech
Sourced in China
The TUNEL apoptosis detection kit is a laboratory tool used to detect and quantify apoptosis, a form of programmed cell death, in samples. The kit utilizes the TUNEL (Terminal deoxynucleotidyl transferase dUTP Nick End Labeling) method to label DNA strand breaks, a hallmark of apoptosis, allowing for the visualization and analysis of apoptotic cells.
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Lab products found in correlation
Eclipse e800, by Nikon (2 mentions)
He staining kit, by Beyotime (1 mentions)
Pas staining kit, by Solarbio (1 mentions)
Ix71 microscope, by Olympus (1 mentions)
Edu staining proliferation kit, by Keygen Biotech (1 mentions)
Inverted microscope, by Nikon (1 mentions)
Transmission electron microscope, by Hitachi (1 mentions)
Anti tumor necrosis factor alpha tnf α antibody, by Abcam (1 mentions)
Neun antibody, by Abcam (1 mentions)
Lsm 710 confocal microscope, by Zeiss (1 mentions)
Annexin 5 fluorescein isothiocyanate fitc apoptosis detection kit, by Keygen Biotech (1 mentions)
Cell cycle detection kit, by Keygen Biotech (1 mentions)
Bca protein assay kit, by Keygen Biotech (1 mentions)
Metformin, by Merck Group (1 mentions)
P akt, by Santa Cruz Biotechnology (1 mentions)
Bcl2 associated x (bax), by Santa Cruz Biotechnology (1 mentions)
P ampk, by Santa Cruz Biotechnology (1 mentions)
Cl caspase3, by Santa Cruz Biotechnology (1 mentions)
P yap, by Santa Cruz Biotechnology (1 mentions)
Hif 1a, by Santa Cruz Biotechnology (1 mentions)
33 protocols using tunel apoptosis detection kit
1
Apoptosis Evaluation in Nude Mouse Tumors
2
Quantifying Apoptosis via TUNEL Assay
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Cell apoptosis was assessed using TUNEL Apoptosis Detection Kit (KeyGen, Nanjing, China), according to the manufacturer's protocol. The nuclei of cells were stained with DAPI. The numbers of TUNEL positive cells (TUNEL+) were counted in randomly selected 10 visual fields through fluorescent microscope. The level of cell apoptosis was calculated by dividing the amount of TUNEL+ cells by that of total cells.
3
Tumor Apoptosis Quantification Protocol
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Implanted tumors were fixed in 4% formalin, paraffinembedded and cut into 4-μm sections before HRP-conjugated dUTP staining. To detected apoptotic cells in the implanted tumors, a TUNEL apoptosis detection kit (Nanjing KeyGen Biotech, KGA7051, China) was used according to the manufacturer’s instructions. All sections were assessed under the microscope (Nikon, Japan). For individual group, the number of apoptotic cells and the total number of cells in five random fields were photographed and counted. The apoptotic index of the cancer cells was calculated using the following formula: Apoptotic index = apoptotic cells/total cells × 100%.
4
Histological Analysis of Tumor Metastasis
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Formalin 10% was used to fix the tissue samples of the tumor and lung, followed by implanting in liquid paraffin. The prepared tissue-embedded blocks were sliced into 5-μm-thick sections. The lung tissue slices were stained with hematoxylin and eosin to visualize the metastatic node. Paraffin was completely removed from the tumor tissue slices, which were then rehydrated and further exposed to autoclaving for 5 min to remove antigenic determinants, followed by incubation in 2.5% hydrogen peroxide for 10–12 min for complete inactivation of peroxidase. The specimens were exposed to goat serum (10%) for 1 h and then incubated at 4°C for 18 h with antibody (Ki67 or CD31). On the next day, the slice was exposed to secondary antibody for 1 h. The slice was then colored using the DAB substrate chromogen system. The TUNEL apoptosis detection kit (KeyGenBioTech, China, Cat. No. KGA7022) was used for detection of TUNEL.
5
TUNEL Assay for Apoptosis Detection
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The TUNEL apoptosis detection kit used in the current study was purchased from KeyGEN Biotech (Nanjing, China). The experimental procedure was performed following the manufacturer’s description. Paraffin-embedded liver sections were incubated in xylene and alcohol with different concentrations. Then, slides were rinsed three times in PBS and further incubated with Proteinase K working solution. The slides interacted with 3% H2O2 at room temperature, then re-incubated with terminal deoxynucleotidyl transferase (TDT) enzyme, streptavidin-horseradish peroxidase (HRP) solution, and 3,3′-diaminobenzidine (DAB) working solution, respectively. Additionally, slides were re-dyed with hematoxylin solution. Following dehydration in alcohol with different proportions, and obtaining transparency using xylene, the slides were sealed with resinene. Images of staining slides were captured by CX33.
6
Histological Assessment of Liver Inflammation
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Mice were anaesthetized using 0.6% pentobarbital sodium (40 mg/kg), and liver tissues were collected and fixed with 4% paraformaldehyde (in PBS) for 48 h at room temperature, dehydrated using an ethanol gradient, cleared with xylene, embedded in paraffin, and cut into 5 μm sections.
To evaluate the degree of inflammatory cell infiltration, liver sections were stained using an H&E staining kit (Beyotime Biotechnology, Shanghai, China). The sections were dewaxed, dehydrated, washed with PBS, and then stained with hematoxylin for 6 min and eosin for 2 min. We evaluated the distribution of liver glycogen using a PAS staining kit (Solarbio, Beijing, China). DNA fragmentation was detected by a TUNEL apoptosis detection kit (KeyGen, Nanjing, China).
To evaluate the degree of inflammatory cell infiltration, liver sections were stained using an H&E staining kit (Beyotime Biotechnology, Shanghai, China). The sections were dewaxed, dehydrated, washed with PBS, and then stained with hematoxylin for 6 min and eosin for 2 min. We evaluated the distribution of liver glycogen using a PAS staining kit (Solarbio, Beijing, China). DNA fragmentation was detected by a TUNEL apoptosis detection kit (KeyGen, Nanjing, China).
7
Apoptosis Detection in Liver Sections
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Apoptotic cells in liver sections were stained by the TUNEL apoptosis detection kit (KeyGEN BioTECH, Nanjing, China). Nuclei were stained with 49,6-diamino-2-phenylindole (DAPI, 1 μg/ml). All images were obtained with an inverted fluorescence microscope (Nikon Eclipse E800, Tokyo, Japan).
8
Visualization of Autophagy and Apoptosis
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After AML12 cells were transfected with GFP-LC3, GFP-LC3 puncta were observed in autophagic cells. To evaluate live tissues, the slides were incubated in an LC3 rabbit monoclonal antibody solution (Cell Signaling, CA, USA) overnight at 4°C and were then incubated in an Alexa Fluor 488 goat anti-rabbit IgG solution (Invitrogen, NY, USA) for 1 h at room temperature. Apoptotic cells were stained with a TUNEL apoptosis detection kit (KeyGEN BioTECH, Nanjing, China). All images were obtained with an inverted fluorescence microscope (Nikon Eclipse E800, Tokyo, Japan).
9
TUNEL Assay for Apoptosis Detection in Rat Hearts
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The apoptotic cells in rat hearts were detected using a TUNEL Apoptosis Detection Kit (KGA7032, KeyGENBioTECH, Nanjing, Jiangsu, China) following the manufacturer's instructions. Briefly, sections were dewaxed, hydrated, permeabilized, blocked using conventional methods and then incubated in the dark with 45 µL equilibration buffer, 1 µL biotin-11-dUTP, and 4 µL TdT for 60 minutes at 37°C, washed 5 minutes in PBS for 3 times, incubated in 0.5 µL streptavidin-HRP in 49.5 µL 1x PBS in the dark at 37°C for 30 minutes, washed 5 minutes in PBS for 3 times, incubated in 2.5 µL DAB-A solution mixed with 50 µL distilled H2O first and then mixed in 2.5 µL of DAB-B and DAB-C solutions at room temperature for 30 seconds to 5 minutes, and washed 3 times in PBS for 5 minutes each and then observed and photographed under a Olympus ix71 microscope.
10
TUNEL Assay for Apoptosis Quantification
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The TUNEL method was used to label apoptotic cells using a TUNEL apoptosis detection kit (KeyGen Biotech Co., Ltd.) according to the manufacturer's protocol. Following treatment with SGI-1027 at a number of concentrations (10, 20 and 30 µmol/l, with 0.1% DMSO as control) for up to 24 h in a humidified atmosphere containing 5% CO2 at 37°C in a 6-well plate, the cells were washed twice with PBS and fixed in 1 ml of 4% paraformaldehyde for 10 min at 4°C, and permeabilized with 0.1% Triton X100 at 25°C for 5 min. The cells washed twice with PBS, stained by the TUNEL mixture for 1 h at 37°C in darkness, and then stained by the Streptavidin-Tetramethylrhodamine for 30 min at 37°C in darkness, followed by staining with DAPI at 37°C for 10 min. The cells were washed with PBS, then mounted and examined using fluorescence microscopy (magnification, ×200) (Olympus IX71; Olympus Corporation, Tokyo, Japan). Apoptotic cells were identified by the condensation and fragmentation of their nucleus. The apoptotic ratio was calculated using the following formula: Apoptotic cell number/seeded cell number ×100%.
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