Cell Death Detection Kit (Sigma-Aldrich) was used to detect cell apoptosis in light of manufacturer’s instructions. DAPI was applied to stain the nucleus of transfected melanoma cells that were cultured in 6-well plates. Fluorescence microscopy (XSP-63B, Shanghai optical instrument factory, Shanghai, China) was adopted to capture the images of stained cells.
Cell death detection kit
Manufactured by Merck Group
Sourced in United Kingdom
The Cell Death Detection Kit is a laboratory product designed to detect and quantify cell death. It provides a reliable and efficient method for the analysis of cell death and apoptosis in research applications.
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Lab products found in correlation
6 protocols using cell death detection kit
1
Apoptosis Detection in Melanoma Cells
2
Ascorbate Induces Apoptosis in Melanoma
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A2058 melanoma cells were seeded in 24 well plates with coverslips and treated with L-ascorbic acid (Sigma-Aldrich St. Louis MO) at different concentrations for 7 days. Glutathione (GSH) was also used to treat cells in a control experiment at a concentration similar to the highest concentration of ascorbate. Apoptotic cells were detected at the end of the treatment utilizing the following three different techniques: (1) colorimetric TUNEL was measured by an in situ apoptosis detection kit (Trevigen, Gaithersburg, MD); (2) fluorescein-based TUNEL was visualized by a cell death detection kit (Sigma-aldrich, St. Louis, MO); and (3) caspase activation was evaluated by incubating cells with CellEvent Caspase-3/7 Green Detection Reagent for 30 min prior to imaging. Each well of TUNEL positive cells or caspase active cells was imaged using a 2D fluorescent microscope system and analyzed with Image J. All experiments were repeated at least 3 times.
3
Assessing BMM Proliferation and Apoptosis
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The cell proliferation reagent, WST-1 (Sigma), was used to detect the proliferation of the BMMs 21 . Each well contained 10 μl of reagent, except for three wells that only contained media (used for subtracting background reactions). After an hour of incubation at 37 °C, a microplate reader was used to interpret its absorbance at 450 nm. The cell death detection kit (Sigma) was utilized to analyze rate of BMM apoptosis in accordance with the manufacturer's protocols.
4
Quantifying Apoptosis in Ovarian Cortex
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A TUNEL (TdT-mediated dUTP-X nick end labeling) assay was performed for detection andquantification of apoptosis within ovaries affected by increasing concentration of BSJ using Cell Death Detection Kit (Sigma, 11684795910), according to the instruction of manufacture with few adjustments. Briefly, the cultured ovaries for 5 days, were fixed, dehydrated, embedded, and sectioned as previously described. Then, the slides were deparaffinized and rehydrated followed by permeabilizing using proteinase K. Next, the labelled sections with TdT were incubated for 2 h in a humidity chamber at 37°C, while the negative controls were omitted the TdT. Subsequently, the sections were counterstained with DAPI. The sections were analyzed immediately using fluorescent imaging (Molecular devices, PICO imageXpress) by calculating the number of apoptotic cells compared to total number of cells in sections of ovarian cortex.
5
Apoptosis Detection in Tissue Samples
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The TUNEL assay was implemented using the Cell Death Detection Kit (Sigma-Aldrich) to detect apoptotic markers on treated tissues. The paraffin-embedded tissues were deparaffinized using xylene and ethanol at the concentration of 50, 70, 85, 95, and 100% and then washed with PBS three times for each step. Apoptosis detection process was followed under the guideline of BD DeadEnd detection kits. The resulting specimens were mounted with ProLong Gold Antifade Mountant with DAPI (4′,6-diamidino-2-phenylindole; P36931, Invitrogen). Images were acquired using a fluorescence microscope (DM6000, Leica). Green fluorescence indicates the location of apoptotic tissue stained by fluorescein isothiocyanate (FITC), and blue fluorescence shows the detected DAPI-stained nuclei.
6
Curcumin Induces Apoptosis in Cells
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Apoptosis was determined through the Cell Death Detection kit (Sigma Aldrich, UK). This ELISA kit detects DNA fragments in the cytoplasm of apoptotic cells. For this experiment, cells were exposed to Cur (5-100 µg/mL) for 24, 48 and 72 h. After each incubation period, cells were lysed and cytoplasmic fractions were obtained. Then, 20 µL of the cytoplasmic fractions were put into a streptavidin-coated 96-well. The mixture of "anti-DNA" and "anti-histone" was added to all wells and additionally incubated for 2 h. After washing the plates, diammonium salt was added and optic densities were measured at 405 nm and 490 nm (Tecan Infinite 200 PRO, Switzerland). The fold changes in each treated well were determined as compared to untreated controls.
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