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Alexa fluor 488 annexin 5 dead cell apoptosis kit with alexa fluor 488 annexin 5 and pi

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The Alexa Fluor® 488 annexin V/Dead Cell Apoptosis Kit with Alexa® Fluor 488 annexin V and PI is a fluorescence-based kit used to detect apoptosis and cell death. It contains Alexa Fluor 488 conjugated annexin V and propidium iodide (PI) for the identification of apoptotic and dead cells.

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Lab products found in correlation

Facsaria 2, by BD (2 mentions) Cellevent caspase 3 7 green flow cytometry assay kit, by Thermo Fisher Scientific (2 mentions) Cytoflex, by Beckman Coulter (2 mentions) Facsaria 2 sorp cell sorter, by BD (1 mentions) Annexin 5 alexa fluor 488, by Thermo Fisher Scientific (1 mentions) Z vad fmk, by Selleck Chemicals (1 mentions)

7 protocols using alexa fluor 488 annexin 5 dead cell apoptosis kit with alexa fluor 488 annexin 5 and pi

1

Evaluating Polymer 1c Cytotoxicity in Hep3B Cells

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EXAMPLE 2

Hep3B cells were seeded onto 6-well plates at a density of 1.6×105 cells per 2 mL DMEM per well. After 24 h, the plating media was replaced with fresh DMEM containing various concentrations of polymers: IC50 and 2×IC50. Cells were harvested after 4, 24, 48 or 72 h incubation, and stained using the Alexa Fluor® 488 annexin V/Dead Cell Apoptosis Kit with Alexa® Fluor 488 annexin V and PI for flow cytometry (Invitrogen, Singapore) according to the manufacturer's instructions. The labeled cells were subjected to flow cytometry analysis (BD FACSAria II, Singapore).

FIGS. 2A-2C depict the percentage of live, apoptotic, and necrotic Hep3B cells after exposure to polymer 1c for 4 hours, in which FIG. 2A is a control, FIG. 2B reflects IC50, and FIG. 2C reflects 2×IC50. FIGS. 3A-3C depict the percentage of live, apoptotic, and necrotic Hep3B cells after exposure to polymer 1c for 72 hours, in which FIG. 3A is a control, FIG. 3B reflects IC50, and FIG. 3C reflects 2×IC50.

US10463746B2. Macromolecular chemotherapeutics (2019-11-05). INTERNATIONAL BUSINESS MACHINES CORPORATION [US], AGENCY FOR SCIENCE, TECHNOLOGY AND RESEARCH [SG]. Inventors: Dylan Boday [US], Wei Cheng [SG], Jeannette M. Garcia [US], James Hedrick [US], Nathaniel H. Park [US], Rudy J. Wojtecki [US], Chuan Yang [SG], YiYan Yang [SG].
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2

Apoptosis Assay for Hep3B Cells

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Example 2

Hep3B cells were seeded onto 6-well plates at a density of 1.6×105 cells per 2 mL DMEM per well. After 24 h, the plating media was replaced with fresh DMEM containing various concentrations of polymers: IC50 and 2×IC50. Cells were harvested after 4, 24, 48 or 72 h incubation, and stained using the Alexa Fluor® 488 annexin V/Dead Cell Apoptosis Kit with Alexa® Fluor 488 annexin V and PI for flow cytometry (Invitrogen, Singapore) according to the manufacturer's instructions. The labeled cells were subjected to flow cytometry analysis (BD FACSAria II, Singapore).

FIGS. 2A-2C depict the percentage of live, apoptotic, and necrotic Hep3B cells after exposure to polymer 1c for 4 hours, in which FIG. 2A is a control, FIG. 2B reflects IC50, and FIG. 2C reflects 2×IC50. FIGS. 3A-3C depict the percentage of live, apoptotic, and necrotic Hep3B cells after exposure to polymer 1c for 72 hours, in which FIG. 3A is a control, FIG. 3B reflects IC50, and FIG. 3C reflects 2×IC50.

US11116844B2. Macromolecular chemotherapeutics (2021-09-14). International Business Machines Corporation [US], Agency for Science, Technology and Research [SG]. Inventors: Dylan Boday [US], Wei Cheng [SG], Jeannette M. Garcia [US], James Hedrick [US], Nathaniel H. Park [US], Rudy J. Wojtecki [US], Chuan Yang [SG], YiYan Yang [SG].
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3

Apoptosis Assay of Cisplatin and Nanoparticles in SKOV-3 Cells

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SKOV-3 cells were seeded at 1×106 cells per well in 6-well plates and further cultured for 24 h. The culture media were replaced by 2 mL of fresh culture media containing 10% FBS. Free cisplatin solution, Zn control, Zn control/siRNAs, NCP-1, NCP-1/siRNAs, NCP-1/siBcl-2, NCP-1/siP-gp, and NCP-1/sisurvivin were added to the cells, respectively, at cisplatin concentration of 5 µM or equivalent nanoparticle concentration of 20 µg/mL. Cells incubated with saline served as control. Following incubation for 24 h, the floating and adherent cells were collected by cell scraper and stained with Alexa Fluor 488 Annexin V/dead cell apoptosis kit with Alexa Fluor 488 annexin V and PI (Invitrogen, USA) according to the manufacturer’s instructions. The apoptosis was examined on a flow cytometer (LSRII Blue, BD, USA).
He C., Liu D, & Lin W. (2014). Self-assembled Nanoscale Coordination Polymers Carrying siRNAs and Cisplatin for Effective Treatment of Resistant Ovarian Cancer. Biomaterials, 36, 124-133.
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4

Quantifying Apoptosis via Annexin V/PI Flow

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Annexin V/propidium iodide (PI) staining was performed to detect apoptotic cells. Seventy-two hours after transfection, 5 × 105 cells were collected and washed twice with ice-cold PBS. The cells were then stained using the Alexa Fluor®488 annexin V/Dead Cell Apoptosis Kit with Alexa® Fluor 488 annexin V and PI for flow cytometry (Invitrogen, CA, USA) according to the manufacturer’s guidelines. Untreated cells served as a negative control for the double staining.
Jiajie T., Yanzhou Y., Hoi-Hung A.C., Zi-Jiang C, & Wai-Yee C. (2017). Conserved miR-10 family represses proliferation and induces apoptosis in ovarian granulosa cells. Scientific Reports, 7, 41304.
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5

Flow Cytometric Immunophenotyping and Apoptosis

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Flow cytometry analysis of the cell surface antigen, CD11b, was performed as described previously [18 (link)]. Briefly, 106 cells were washed in PBS, blocked with human IgG (Sigma-Aldrich), then incubated with Alexa Fluor 700 Mouse Anti-Human-CD11b for 30 min. The proportion of live/dead/apoptotic cells was assessed with the Alexa Fluor® 488 annexin V/Dead Cell Apoptosis Kit with Alexa Fluor® 488 annexin V and PI (Thermo Fisher). Briefly, 5 × 105 cells were washed and incubated with Alexa Fluor® 488 annexin V and 100 μg/mL PI for 15 min. Fluorescence was measured with a FACSAria II SORP cell sorter (BD Biosciences). Analysis was performed in FlowJo v10.
Jacobson E.C., Perry J.K., Long D.S., Olins A.L., Olins D.E., Wright B.E., Vickers M.H, & O’Sullivan J.M. (2018). Migration through a small pore disrupts inactive chromatin organization in neutrophil-like cells. BMC Biology, 16, 142.
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6

Evaluating Apoptotic Mechanisms in SKOV3 and A2780CIS Cells

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SKOV3 and A2780CIS cells were plated in T25 flasks 24 h before drug exposure. Afterward, samples were treated with 37°C warm media mixed with 100 nM onalespib, 500 nM cisplatin, or a combination of onalespib and cisplatin for 96 h before flow cytometry. Harvested cells were washed in cold PBS and stained with propidium iodide (PI) and Alexa Fluor 488 annexin V (Alexa Fluor®488 Annexin V/Dead Cell Apoptosis Kit with Alexa Fluor 488 annexin V and PI for flow cytometry, ThermoFisher Scientific, Sweden) according to manufactures instructions. CellEventTM Caspase-3/7 Green Flow Cytometry Assay Kit (Thermo Fisher Scientific, Sweden) was used to analyze caspase 3/7 activity. Caspase activity inhibition on SKOV3 and A2780CIS apoptosis were evaluated by pan-caspase inhibitor z-VAD-FMK (Selleckchem, Germany). Cells were pretreated with or without 20 μM z-VAD-FMK for 1 h followed by incubation with 500 nM cisplatin and 100 nM onalespib. Apoptotic cells were visualized using a CytoFLEX (Beckman Coulter, Krefeld, Germany). Obtained data were analyzed by FlowJoTM Software for Windows (Version 10.6.1. Becton, Dickinson and Company, Ashland, OR, United States). One-way ANOVA followed by Tukey’s multiple comparison’s test determined significance, where p < 0.05 was considered significant. The number of replicates within each experimental group was two. Each experiment was repeated three times.
Mortensen A.C., Mohajershojai T., Hariri M., Pettersson M, & Spiegelberg D. (2020). Overcoming Limitations of Cisplatin Therapy by Additional Treatment With the HSP90 Inhibitor Onalespib. Frontiers in Oncology, 10, 532285.
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7

Quantifying Apoptosis in BHT-101 Cells

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After exposure to sorafenib (15 μM, onalespib (100 nM), or the combination thereof BHT-101 cells were harvested and washed in PBS and stained with propidium iodide (PI) and Alexa Fluor 488 annexin V (Alexa Fluor®488 Annexin V/Dead Cell Apoptosis Kit with Alexa Fluor 488 annexin V and PI for flow cytometry, ThermoFisher Scientific, Sweden) according to manufactures instructions. CellEventTM Caspase-3/7 Green Flow Cytometry Assay Kit (Thermo Fisher Scientific, Sweden) was used to analyze caspase 3/7 activity. Apoptotic cells were visualized using a CytoFLEX (Beckman Coulter, Krefeld, Germany). Obtained data were analyzed by FlowJoTM Software for Windows (Version 10.9). The number of replicates within each experimental group was four. Each experiment was repeated two times.
Mortensen A.C., Berglund H., Hariri M., Papalanis E., Malmberg C, & Spiegelberg D. (2023). Combination therapy of tyrosine kinase inhibitor sorafenib with the HSP90 inhibitor onalespib as a novel treatment regimen for thyroid cancer. Scientific Reports, 13, 16844.
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