Hoechst 33258, bicinchoninic acid (BCA) protein assay kit, enhanced chemiluminescence (ECL) solution, and radioimmunoprecipitation assay (RIPA) buffer were supplied by Beyotime Institute of Biotechnology (Haimen, China). The cell counting kit-8 (CCK-8) was obtained from Dojindo Molecular Technologies, Inc. (Kumamoto, Japan). F12/Dulbecco's modified Eagle's medium (DMEM-F12) and fetal bovine serum (FBS) were purchased from HyClone (GE Healthcare Life Sciences, Logan, UT, USA). Primary antibodies for caspase-3 (cat. no. 9662) and GAPDH (cat. no. 5174) were purchased from Cell Signaling Technology, Inc. (Danvers, MA, USA), and horseradish peroxidase (HRP)-conjugated secondary antibodies (cat. no. 00001-9) were obtained from ProteinTech Group, Inc. (Danvers, MA, USA). Annexin V/propidium iodide (PI) apoptosis kit was supplied by BD Pharmingen (BD Biosciences, San Jose, CA, USA). Lipofectamine RNAiMAX and Opti-MEM medium were supplied by Invitrogen (Thermo Fisher Scientific, Inc., Waltham, MA, USA). The miRcute miRNA Isolation kit, miRcute miRNA First-strand cDNA Synthesis kit and miRcute miRNA quantitative polymerase chain reaction (qPCR) detection kit were obtained from Tiangen Biotech Co., Ltd. (Beijing, China).
Annexin 5 propidium iodide apoptosis kit
Manufactured by BD
Sourced in United States
The Annexin V/propidium iodide (PI) apoptosis kit is a laboratory tool used to detect and quantify apoptosis, a form of programmed cell death, in cell samples. The kit contains Annexin V, a protein that binds to phosphatidylserine, and propidium iodide, a dye that stains the nucleic acids of cells with compromised cell membranes. This combination allows for the identification and differentiation of viable, early apoptotic, late apoptotic, and necrotic cells through flow cytometry or fluorescence microscopy analysis.
Automatically generated - may contain errors
Lab products found in correlation
Enhanced chemiluminescence, by Beyotime (1 mentions)
Cell counting kit 8 cck 8, by Dojindo Laboratories (1 mentions)
Lipofectamine rnaimax, by Thermo Fisher Scientific (1 mentions)
Opti mem medium, by Thermo Fisher Scientific (1 mentions)
Bca protein assay kit, by Beyotime (1 mentions)
Hoechst 33258, by Beyotime (1 mentions)
Mircute mirna isolation kit, by Tiangen Biotech (1 mentions)
Mircute mirna first strand cdna synthesis kit, by Tiangen Biotech (1 mentions)
Annexin 5 binding buffer, by BD (1 mentions)
Cellquest software, by BD (1 mentions)
Facscalibur instrument, by BD (1 mentions)
Facscalibur flow cytometer, by BD (1 mentions)
Facscan flow cytometer, by BD (1 mentions)
Flow cytometry, by BD (1 mentions)
Paint a gate software, by BD (1 mentions)
8 protocols using annexin 5 propidium iodide apoptosis kit
1
Apoptosis Assay Protocol for Cell Lines
2
Synergistic and Sensitizing Effects of Curcumin and Cisplatin on NSCLC Cells
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The apoptosis assay was conducted using the Annexin V/propidium iodide (PI) apoptosis kit purchased from BD Pharmingen. In brief, 9.0×105 cells/well of sorted and unsorted NSCLC cells were seeded into 6-well plates and incubated overnight. Direct combination (synergistic effects) of both curcumin and cisplatin on the NSCLC cell lines was performed by incubation of the cells in medium containing the single treatment (cisplatin or curcumin) and combination of both using the IC50 doses for 48 h. Indirect combination (sensitising effects) of curcumin was performed by incubating the NSCLC cell lines with curcumin (IC50 value) for 24 h, followed by incubation with low dose cisplatin (3 µM) for another 24 h. After treatments for 48 h (synergistic and sensitisation), both NSCLC cell lines were harvested by trysinisation and collected by centrifugation. The cell pellet was suspended in 100 µl of 1X Annexin V binding buffer (Becton Dickinson BD) and 1 µl of Annexin V-FITC was added. Antibody incubation was performed at 4°C for 20 min, and 1 µl of PI was later added before FACS acquisition. Stained cells were subjected to flow cytometric analysis using a FACSCalibur instrument (Becton Dickinson BD), and a total of 10,000 events were acquired and analyzed using Cell Quest software (Becton Dickinson BD).
3
Annexin V-FITC/PI Apoptosis Assay
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Cells treated with different concentrations of GXN and ISO were processed and collected into centrifuge tubes and subjected to flow cytometry using the Annexin V/Propidium Iodide (PI) Apoptosis Kit (BD Bioscience, USA). Cells were washed 3 times with PBS, then resuspended in 1× binding buffer. After that, cells were incubated with 5 μl Annexin V-FITC and 10 μl propidium iodide, each for 15 min in the dark (Wang et al., 2009) (link). 300 μl of 1× Binding buffer was added and mixed, then the cell suspension was transferred into a 5 ml flow cytometer under dark conditions. Samples were detected by FACSCalibur flow cytometer (BD Bioscience, San Jose, CA, USA) within an hour. Data were processed using FlowJo 10 (FlowJo, LLC, Ashland, OR, USA).
4
SPINK13 Regulates Cell Proliferation and Apoptosis
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Cells were seeded in 96-well plates at a density of 5 × 10 3 cells per well. After 24 h, the cells were transfected with 50 nM of SPINK13-overexpressing vector (pcDNA-SPINK13), control vector (pcDNA3.1), siRNA-SPINK13 (si-SPINK13) or siRNA-NC (si-NC) and allowed to grow for another 48 h. Cell proliferation was measured using the MTT assay as reported previously [18] . The OD value was represented as the viability index. Three independent experiments were performed with quadruplicate samples.
The Annexin V/propidium iodide (PI) apoptosis kit (BD Biosciences, San Jose, CA, USA) was employed to evaluate cellular apoptosis. The cells were harvested 48 h after tranfection with plasmids or siRNAs and then stained with Annexin V/PI following the manufacturer's protocol. The results were analysed using a FACScan flow cytometer (BD Biosciences, San Diego, CA, USA).
The Annexin V/propidium iodide (PI) apoptosis kit (BD Biosciences, San Jose, CA, USA) was employed to evaluate cellular apoptosis. The cells were harvested 48 h after tranfection with plasmids or siRNAs and then stained with Annexin V/PI following the manufacturer's protocol. The results were analysed using a FACScan flow cytometer (BD Biosciences, San Diego, CA, USA).
5
Annexin V-based Cell Apoptosis Assay
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A cell apoptosis assay was performed using an Annexin V/propidium iodide apoptosis kit (BD Biosciences, Franklin Lakes, NJ, USA) following the manufacturer’s protocol. Adherent cells were cultured in serum-free medium for 24 h. The cells were detached with 0.25% trypsin (without ethylenediaminetetraacetic acid), washed, re-suspended with binding buffer, and stained with annexin V-fluorescein isothiocyanate and propidium iodide. The percentages of Annexin V-positive and propidium iodide-negative cells were determined by flow cytometry using a FACS Cater -plus flow cytometer (BD Biosciences).
6
Annexin V/PI Apoptosis Assay by Flow Cytometry
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Apoptosis (early and late stage) was determined using an Annexin V/propidium iodide apoptosis kit and flow cytometry (BD Biosciences, USA) with Cell Quest software. Raji, Daudi, Jurkat and CEM cells, after various treatments for 24, 48 and 72 h, were rinsed twice with cold PBS and resuspended in 195 μl of binding buffer solution. Cells were stained with 5 μl FITC-labeled Annexin V and 10 μl propidium iodide for 20 min at room temperature in the dark. Cells treated with medium alone were used as a control.
7
Annexin V-FITC/PI Apoptosis Assay
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Apoptosis (early and late stage) was determined using an Annexin V/propidium iodide apoptosis kit and flow cytometry (BD Biosciences, San Jose, CA) with Cell Quest software. Raji, Daudi, Jurkat, and CEM cells, after various treatments for 24, 48, and 72 h, were rinsed twice with cold PBS and resuspended in 195 μL of binding buffer solution. Cells were stained with 5 μl FITC-labeled Annexin V and 10 μL propidium iodide for 20 min at room temperature in the dark. Cells treated with medium alone were used as a control (Mohamed Subarkhan et al., 2016 ; Subarkhan and Ramesh, 2016 ; Chung et al., 2017 (link); Mohan et al., 2018 ; Mohamed Subarkhan et al., 2019 (link); Sathiya Kamatchi et al., 2020 (link)).
8
Quantification of Cell Apoptosis
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To quantitate the amount of apoptosis, after harvesting cells which were incubated by various textures in complete medium, cells were washed twice with washing buffer (phosphate buffered saline 0.15 M, 0.5% bovine serum albumin, 0.1% NaN3). Fresh B-cells were resuspended in 100 µl washing buffer and stained with appropriate fluorochrome-conjugated mAbs and incubated for 45 min at 4°C in the dark atmosphere to evaluate the amount of apoptosis by using Annexin V/Propidium iodide apoptosis kit (BD Biosciences, San Jose, CA, USA). Then 1000 cells/sample was acquired in an FAC Scan flow cytometer (Partec, Nuremberg, Germany). Moreover, the proportions of labeled cells were analyzed using Paint-A-Gate software (Becton and Dickinson, Sunnyvale, USA). Data analysis was performed using the Flomax software (Partec, Nuremberg, Germany).
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