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Apoalert annexin 5 fitc apoptosis kit

Manufactured by Takara Bio
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The ApoAlert™ Annexin V-FITC Apoptosis Kit is a laboratory product that detects apoptosis, a type of programmed cell death, in cell samples. The kit utilizes Annexin V, a protein that binds to phosphatidylserine, a molecule that becomes exposed on the cell surface during apoptosis. The Annexin V is conjugated with the FITC fluorescent dye, allowing for the visualization and quantification of apoptotic cells.

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Lab products found in correlation

Lsrfortessa x 20, by BD (3 mentions) Facscelesta, by BD (3 mentions) Facscalibur, by BD (2 mentions) Cell proliferation kit 1 mtt, by Merck Group (1 mentions) Anti cleaved caspase 3, by Cell Signaling Technology (1 mentions) Planapon objective lens, by Olympus (1 mentions) Ix83 inverted microscope, by Olympus (1 mentions) Cellsens, by Olympus (1 mentions) Trypan blue, by Thermo Fisher Scientific (1 mentions) Facscanto 2, by BD (1 mentions) Fluorescence activated cell sorting (facs), by BD (1 mentions) Lrs fortessa, by BD (1 mentions) Guava easycyte flow cytometer, by Merck Group (1 mentions) Zymolyase 20t, by Nacalai Tesque (1 mentions) Hepes, by Merck Group (1 mentions)

Spelling variants of this product

Annexin V-FITC Apoptosis Kit (9 citations) ApoAlert (7 citations) Annexin V-FITC (6 citations) ApoAlert Annexin V Kit (4 citations) APOAlert® Annexin V (3 citations) Apoalert Annexin V Apoptosis Kit (2 citations)

18 protocols using apoalert annexin 5 fitc apoptosis kit

1

Detecting Apoptotic Tumor Cells by Flow Cytometry

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To detect apoptotic tumor cells, siRNA-treated cells were stained using the ApoAlert Annexin V-FITC Apoptosis Kit (Takara Bio Inc, Shiga, Japan) according to the manufacturer’s instructions. The staining pattern was analyzed by flow cytometry.
Shinada M., Kato D., Kamoto S., Yoshimoto S., Tsuboi M., Yoshitake R., Eto S., Ikeda N., Saeki K., Hashimoto Y., Takahashi Y., Chambers J., Uchida K., Kaneko M.K., Fujita N., Nishimura R., Kato Y, & Nakagawa T. (2020). PDPN Is Expressed in Various Types of Canine Tumors and Its Silencing Induces Apoptosis and Cell Cycle Arrest in Canine Malignant Melanoma. Cells, 9(5), 1136.
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2

Annexin V-FITC Apoptosis Assay

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The ApoAlert™ Annexin V-FITC Apoptosis Kit (Takara) was used according to the manufacturer’s instructions. Cells were seeded and stimulated for 72 hr. Analysis was carried out on a FACS Calibur (BD Biosciences). Quadrants were defined by single stain controls and the percentages of apoptotic cells were calculated from both early and late apoptotic fractions. A 2-way ANOVA test with Tukey correction was carried out for statistical analysis.
Brown K., Jenkins L.M., Crooks D.R., Surman D.R., Mazur S.J., Xu Y., Arimilli B.S., Yang Y., Lane A.N., Fan T.W., Schrump D.S., Linehan W.M., Ripley R.T, & Appella E. (2023). Targeting mutant p53-R248W reactivates WT p53 function and alters the onco-metabolic profile. Frontiers in Oncology, 12, 1094210.
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3

Annexin V-FITC Apoptosis Detection

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The Annexin V-FITC apoptosis detection kit (ApoAlert Annexin V-FITC Apoptosis Kit, TaKaRa) was used to detect apoptosis by flow cytometry. Cells were seeded at 6-well plates at concentration of 5 × 105 cells/well, cultured for 18 h, and then the tested agent was applied for the indicated periods. Afterward, cells were washed with PBS, resuspended in binding buffer, and anti-Annexin V FITC-conjugated antibody and propidium iodide were added to 100 µl aliquots. The mixtures were incubated for 15 min at room temperature, supplemented with binding buffer to 500 µl, and processed using BD FACSCalibur (BD Biosciences). Data were analyzed in Flowing Software version 2.5.1 (Flowing Software, http://www.uskonaskel.fi/flowingsoftware).
Pilžys T., Marcinkowski M., Kukwa W., Garbicz D., Dylewska M., Ferenc K., Mieczkowski A., Kukwa A., Migacz E., Wołosz D., Mielecki D., Klungland A., Piwowarski J., Poznański J, & Grzesiuk E. (2019). ALKBH overexpression in head and neck cancer: potential target for novel anticancer therapy. Scientific Reports, 9, 13249.
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4

Apoptosis Evaluation via Annexin V and PI

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For annexin V and propidium iodide (PI) staining, cells were stained using the ApoAlert Annexin V‐FITC Apoptosis Kit (Takara Bio), according to the manufacturer’s instructions. For cleaved caspase‐3 and cytochrome c immunostaining, cells were fixed with 3% paraformaldehyde in PBS for 10 minutes at 37°C, and permeabilized with 1% Triton X‐100 in PBS for 5 minutes. Fixed cells were incubated with anti‐cleaved caspase‐3 (1:2000, 9664; Cell Signaling Technology) and anti‐cytochrome c (1:2000, NBP2‐24872; NovusBio) Abs for 1 hour, followed by incubation with secondary Abs coupled with Alexa Fluor 488/594 (Thermo Fisher Scientific, 1:3000) for 1 hour. Then cells were observed under an Olympus IX‐83 inverted microscope controlled by cellSens (Olympus) using a ×60 1.42 NA PlanApoN objective lens (Olympus). For the MTT assay, 1 × 103 cells were seeded onto 96‐well culture plates and were analyzed using a Cell Proliferation Kit I (MTT) (Merck), according to the manufacturer’s instructions.
Hino M., Iemura K., Ikeda M., Itoh G, & Tanaka K. (2021). Chromosome alignment‐maintaining phosphoprotein CHAMP1 plays a role in cell survival through regulating Mcl‐1 expression. Cancer Science, 112(9), 3711-3721.
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5

Yeast Cell Viability Assays

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Cells were grown in SMD media until A600 reached approximately 1. For colony-forming unit assays, cells were normalized to A600 = 0.5/ml, and 200 μl of each culture was collected into 96-well plates. Serial dilutions were done using multichannel pipettor, and 10 μl were plated onto SMD solid media. Plates were incubated for 2 days at 30 °C, and number of colonies were counted. Annexin V/PI co-staining assays using ApoAlert Annexin V-FITC Apoptosis kit (Takara, 630109) were performed as described previously (92 (link)). For trypan blue staining, 10 μl of cell suspension in PBS were mixed with 0.4% trypan blue (Gibco, 15250061) for 2 to 3 min at room temperature, and number of cells stained with the dye were counted.
Pal A., Paripati A.K., Deolal P., Chatterjee A., Prasad P.R., Adla P, & Sepuri N.B. (2022). Eisosome protein Pil1 regulates mitochondrial morphology, mitophagy, and cell death in Saccharomyces cerevisiae. The Journal of Biological Chemistry, 298(11), 102533.
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6

Annexin V Apoptosis Assay by Flow Cytometry

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Cells (1 × 106) were collected with trypsin-EDTA, washed twice with PBS, and then fixed in 70% ethanol for 24 h at 20°C. The cells were then centrifuged and washed with 200 μL of 1× binding buffer from the ApoAlert Annexin V–FITC Apoptosis kit (cat #630109; Clontech, Mountain View, CA, USA). The cells were incubated for 5–15 min in the dark with 5 μL of Annexin V solution from the kit. After staining, the cells were washed with PBS. The cell-cycle distribution was determined by flow cytometry on a Guava easyCyte (Luminex, Austin, TX, USA) and analyzed using the FlowJo software.
Chei S., Oh H.J., Song J.H., Seo Y.J., Lee K, & Lee B.Y. (2019). Magnolol Suppresses TGF-β-Induced Epithelial-to-Mesenchymal Transition in Human Colorectal Cancer Cells. Frontiers in Oncology, 9, 752.
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7

Cell Cycle and Apoptosis Analysis in Breast Cancer Cells

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For cell cycle assays, MDA-MB-231, MDA-MB-468, MCF7 and ZR75 cells were treated with vehicle or 20 μM Imipramine or 40 μM imipramine for 96 h and then fixed with 70% ethanol for at least 24 h before staining with propidium iodide, as previously described [30 ]. Cell cycle distribution was determined using a BD FACSCelesta™ or BD LSRFortessa™ X-20 instruments. For apoptosis assays, cells were harvested after 96 h treatment with Vehicle or 20 μM Imipramine or 40 μM imipramine and then stained with annexin V/propidium iodide mixture using the ApoAlert™ Annexin V-FITC Apoptosis Kit (Clontech, Cat # 630110). The percent of annexin V-propidium iodide-positive cells were determined using flow cytometry as described previously [30 ]. The samples were analyzed using a BD FACSCelesta™ or BD LSRFortessa™ X-20 instrument. Data were processed using BD FACSDiva™ or FlowJo v10.7.2 software and visualized using a GraphPad Prism program.
Timilsina S., Rajamanickam S., Rao A., Subbarayalu P., Nirzhor S., Abdelfattah N., Viswanadhapalli S., Chen Y., Jatoi I., Brenner A., Rao M.K., Vadlamudi R, & Kaklamani V. (2022). The antidepressant imipramine inhibits breast cancer growth by targeting estrogen receptor signaling and DNA repair events. Cancer letters, 540, 215717.
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8

Apoptosis Assay for B16F10 Cells

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2 × 105 B16F10 cells were plated in each well of 6-well plate and allowed to grow for 24 h after which they were treated with PTX drug, lipo-PTX/Scrambled siRNA, lipo- Bcl-2 siRNA and lipo-PTX/Bcl-2 siRNA and allowed to incubate for 24 h and 48 h. ApoAlert™ Annexin V-FITC Apoptosis Kit was used to measure apoptotic cells by flow cytometry according to the manufacturer’s instruction (Clontech Laboratories, Inc. CA, USA). Cells collected by trypsinization were washed twice with ice cold Dulbecco’s phosphate buffered saline (DPBS) and resuspended in 200 μL of 1X binding buffer containing 5 μL Annexin V and 10 μL Propidium iodide for 15 min at room temperature in the dark. Apoptosis of cells was measured on a Beckman Coulter flow cytometer (DakoCytomation, Inc. CA, USA). At least 20,000 gated events were acquired from each sample. Results are expressed as the percentage of apoptotic cells (PI and Annexin V positive) in the gated cell population.
Reddy T.L., Garikapati K.R., Reddy S.G., Reddy B.V., Yadav J.S., Bhadra U, & Bhadra M.P. (2016). Simultaneous delivery of Paclitaxel and Bcl-2 siRNA via pH-Sensitive liposomal nanocarrier for the synergistic treatment of melanoma. Scientific Reports, 6, 35223.
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9

Analyzing Hepatic Macrophage Apoptosis

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Liver non‐parenchymal mononuclear cells were isolated from the liver and subjected to flow cytometry, as previously described.17 The involvement of CHI3L1 in hepatic F4/80+ macrophage apoptotic functions was investigated by examining the expression of annexin V and propidium iodide (PI) by flow cytometry (BD FACSCanto II; Becton Dickinson, Franklin Lakes, NJ, USA) with an ApoAlert Annexin V‐FITC Apoptosis Kit (Clontech Laboratories, Mountain View, CA, USA), following the manufacturer's protocol.
Higashiyama M., Tomita K., Sugihara N., Nakashima H., Furuhashi H., Nishikawa M., Inaba K., Wada A., Horiuchi K., Hanawa Y., Shibuya N., Kurihara C., Okada Y., Nishii S., Mizoguchi A., Hozumi H., Watanabe C., Komoto S., Yamamoto J., Seki S., Miura S, & Hokari R. (2019). Chitinase 3‐like 1 deficiency ameliorates liver fibrosis by promoting hepatic macrophage apoptosis. Hepatology Research, 49(11), 1316-1328.
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10

Quantifying Apoptosis and Nuclear Damage

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Apoptosis and cell nuclear damage were determined by Annexin V- FITC and PI dual cell staining; stained cells were subsequently analyzed by flow cytometry [60 (link)]. Cells grown in 100 mm dishes were transduced with dE1 or dE1/DCN at an MOI of 20–50, or with dB or dB/DCN at an MOI of 0.5–5. As a positive control for the induction of apoptosis, cells were treated with CPT at 1 mM. After two days of treatment, cells were processed with the ApoAlert Annexin V-FITC apoptosis kit (Clontech, Palo Alto, CA, USA) according to the manufacturer's instructions. Apoptosis and nuclear cell damage were quantified on a fluorescence-activated cell sorter (FACS; Becton Dickinson, Sunnyvale, CA, USA) and data from 10,000 events were collected for further analysis.
Yoon A.R., Hong J, & Yun C.O. (2017). Adenovirus-mediated decorin expression induces cancer cell death through activation of p53 and mitochondrial apoptosis. Oncotarget, 8(44), 76666-76685.
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