Cells from different groups were trypsinized and resuspended with cold PBS. An Annexin V-fluorescein isothiocyanate (FITC) apoptosis kit (BD Biosciences, Franklin Lakes, NJ, USA) was used to detect phosphatidylserine externalization as an index of apoptosis. The cells were washed and incubated for 15 min at room temperature in the presence of Annexin V labeled with FITC and propidium iodide (PI). 10,000 cells were loaded and excited at 488 nm, and emission was measured at 530 and 584 nm to assess FITC and PI fluorescence, respectively. Analysis was conducted with BD FACS Calibur flow cytometer (BD Biosciences, Franklin Lakes, NJ, USA). The number of gated cells was plotted on a dot plot with reference to both Annexin V and PI staining.
Annexin 5 fluorescein isothiocyanate fitc apoptosis kit
Manufactured by BD
Sourced in United States
The Annexin V-fluorescein isothiocyanate (FITC) apoptosis kit is a laboratory assay used to detect and quantify apoptosis, a form of programmed cell death. The kit utilizes Annexin V, a calcium-dependent phospholipid-binding protein, which has a high affinity for phosphatidylserine (PS). During apoptosis, PS is translocated from the inner to the outer leaflet of the plasma membrane, where it can be detected by Annexin V conjugated to the fluorescent dye FITC.
Automatically generated - may contain errors
Lab products found in correlation
Accuri c6, by BD (4 mentions)
Facscalibur flow cytometer, by BD (3 mentions)
Biosciences accuri c6, by BD (3 mentions)
10 n nonyl acridine orange a1372, by Thermo Fisher Scientific (2 mentions)
Tetramethylrhodamine ethyl ester tmre t669, by Thermo Fisher Scientific (2 mentions)
Rnase a, by Merck Group (2 mentions)
Propidium iodide, by Merck Group (2 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Phosphorylated p38, by Abcam (1 mentions)
C jun n terminal kinases, by Abcam (1 mentions)
P jnk, by Abcam (1 mentions)
P erk, by Abcam (1 mentions)
Macsquant, by Miltenyi Biotec (1 mentions)
Cellquest 6, by BD (1 mentions)
Anti β actin ab8226, by Abcam (1 mentions)
Puromycin p8833, by Merck Group (1 mentions)
5 and 6 chloromethyl 2 7 dichlorodihydrofluorescein diacetate cm h2dcfda, by Thermo Fisher Scientific (1 mentions)
Facsaria fusion flow cytometer, by BD (1 mentions)
Cellquest software, by BD (1 mentions)
β actin ab8226, by Abcam (1 mentions)
23 protocols using annexin 5 fluorescein isothiocyanate fitc apoptosis kit
1
Apoptosis Detection by Annexin V-FITC
2
Role of PCSK9 in Ox-LDL-Induced Endothelial Cell Apoptosis
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The human umbilical vein endothelial cell (HUVEC) line EAhy926 was obtained from the Institute of Pharmacology, Medical University of Tianjin (Tianjin, China). Ox-LDL was purchased from Xinyuan Jiahe Biotechnology (Beijing, China). Basic Dulbecco's modified Eagle's medium (DMEM) and fetal bovine serum were purchased from Invitrogen (Thermo Fisher Scientific, Inc., Waltham, MA, USA). RNase inhibitor and Moloney murine leukemia virus reverse transcriptase was purchased from Takara Biotechnology Co., Ltd. (Dalian, China). Monoclonal antibodies against PCSK9, B-cell lymphoma 2 (Bcl-2), bcl-2-like protein 4 (Bax), caspase-3, p38, phosphorylated (p)-p38, extracellular signal-regulated kinases (ERK), p-ERK, c-Jun N-terminal kinases (JNK) and p-JNK were purchased from Abcam (Cambridge, MA, USA). SYBR Green PCR Premix was purchased from Biocentury TransGene (Beijing, China). Annexin V fluorescein isothiocyanate (FITC) apoptosis kit was purchased from BD Biosciences (Franklin Lakes, NJ, USA). The flow cytometer (version, BDFACS Verse) was purchased from BD Biosciences and FlowJo software version 7.6 (FlowJo, LLC, Ashland, OR, USA) was used. The lentiviral packaging system, containing helper plasmids and target plasmids, was purchased from CWBIO (Beijing, China).
3
Detecting BMEC Apoptosis by Flow Cytometry
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To detect the apoptosis of BMECs, flow cytometry was performed after 24-hour OGD treatment, as previously described (Bai et al., 2018). Briefly, the cells were collected and resuspended in 500 µL PBS, fixed, and stained with an Annexin V fluorescein isothiocyanate (FITC) apoptosis kit (BD Biosciences, New Jersey, USA). The apoptotic and non-apoptotic cells were separated using a Beckman CytoFLEX (BD Biosciences).
4
Bupivacaine-Induced Apoptosis Quantification
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Cell death by apoptosis was analyzed using the Annexin V-fluorescein isothiocyanate (FITC) apoptosis kit (BD Biosciences). Tumor cells were treated with either 0.25% or 0.5% bupivacaine for various durations. Initially, 1x105 cells were seeded in 6-well plates, and when confluence reached 80%, they were exposed to either 0.25% or 0.5% bupivacaine. After 24 h, both floating and attached cells were harvested and then washed. Flow cytometry with FITC-conjugated Annexin-V/propidium iodide (PI) double staining was used to assess the number of apoptotic cells. Samples were analyzed by flow cytometry (MACSQuant; Miltenyi Biotec). Using the FlowJo software (v10; TreeStar), measurements were displayed as 4 quadrants, in which the lower right quadrant represented the apoptosis rate during the early stages, the upper right quadrant indicated advanced apoptosis rate, the upper left quadrant represented dead cells, and the lower left quadrant represented living cells. The apoptotic rate was calculated as early apoptosis (Ann+/PI-) and late apoptosis (Ann+/PI+). This experiment was repeated 3 times.
5
Apoptosis Analysis of Cancer Cells
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The apoptosis of MGC-803 and BGC-823 cells following different treatments was analyzed using an Annexin V-fluorescein isothiocyanate (FITC) apoptosis kit (BD Biosciences) by flow cytometry. Cells were collected and stained with Annexin V-FITC and propidium iodide solutions. Subsequently, the apoptosis of the cells was examined using a FACSCalibur flow cytometer (BD Biosciences) with CellQuest 6.0 software (BD Biosciences, Franklin Lakes, NJ, USA).
6
Mitochondrial Oxidative Stress Regulation
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Reagents utilized in this study include diclofenac sodium (D6899) and puromycin (p8833) from Sigma Aldrich (St. Louis, MO, USA); the annexin V-fluorescein isothiocyanate (FITC) apoptosis kit (556547) was purchased from BD biosciences (San Jose, CA, USA); acetyl-Leu-Glu-His-Asp-7-amino-4-trifluoromethylcoumarin (Ac-LEHD-AFC) and acetyl-Asp-GluVal-Asp-7-amino-4-methylcoumarin (Ac-DEVD-AMC) were from BIOMOL (Hamburg Germany); 5-(and-6)-chloromethyl-2′,7′-dichlorodihydrofluorescein diacetate (CM-H2DCFDA), 10-N-nonyl-acridine orange (NAO) (A1372), and tetramethylrhodamine ethyl ester (TMRE) (T669) were from Molecular Probes ( Eugene, OR, USA); mitochondria peroxy-yellow-1 (MitoPY1) (4428) was from Tocris Biosciences; FuGene6 (E2311) and pSUPER-puro vector were from Promega (Madison, WI, USA) and OligoEngine (Seattle, WA, USA) respectively. The following antibodies were used: anti-PrxIII (LF-MA0329) and anti-β-actin (ab8226) were purchased from Abcam (Cambridge, UK); anti-catalase (LF-PA0060) was from Abfrontier (Seoul, Republic of Korea). Adenovirus expressing human catalase with a mitochondrial leader sequence (mito-Catalase) was used as described previously [14 (link),15 (link)].
7
Annexin V Apoptosis Assay by Flow Cytometry
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Cell apoptosis was evaluated by flow cytometry assay using the Annexin V-fluorescein isothiocyanate (FITC) apoptosis kit (BD Biosciences, San Jose, CA, USA). The prepared cells were double-labeled with Annexin V-FITC (5 μL) and propidium iodide (5 μL) for 15 min following the manufacturer’s protocol. The apoptosis rate was immediately analyzed in a FACSAria™ Fusion flow cytometer using CellQuest software (BD Biosciences, San Jose, CA, USA).
8
Annexin V-FITC Apoptosis Assay
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Cell apoptosis was analyzed using an Annexin V-fluorescein isothiocyanate (FITC) Apoptosis kit (BD Biosciences) in accordance with the manufacturer's protocol. Transfected SK-BR-3 and BT-549 cells were stained with Annexin V-FITC and propidium iodide (PI) solutions at 4°C for 30 min. Then, the cell apoptosis rate was detected by a FACSCalibur flow cytometer (Becton, Dickinson and Company) and analyzed using BD Accuri C6 software (version 1.0; Becton, Dickinson and Company).
9
Apoptosis Pathway Reagents Protocol
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The following reagents were used in this study: cytochrome C (556433) and the annexin V-fluorescein isothiocyanate (FITC) apoptosis kit (556547) were purchased from BD biosciences; caspase-3 (9662), caspase-9 (9508), and poly(ADP-ribose)polymerase (PARP) (9542) were from Cell Signaling Technology; Prx-SO2 (ab16830), PrxI (ab15571), and β-actin (ab8226) were from Abcam; PrxV (LF-MA0002) was from Invitrogen; 10-N-nonyl-acridine orange (NAO) (A1372) and tetramethylrhodamine ethyl ester (TMRE) (T669) were from Molecular Probes; peroxy-orange-1 (PO-1) (4944) and mitochondria peroxy-yellow-1 (MitoPY) (4428) were from Tocris Biosciences; puromycin (p8833), FuGene6 (E2311), and pSUPER-puro vector were from Sigma Aldrich, Promega and OligoEngine, respectively.
10
Annexin V-FITC Apoptosis Assay
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Cell apoptosis was analyzed using an Annexin V-fluorescein isothiocyanate (FITC) Apoptosis kit (BD Biosciences, San Diego, CA, USA) following the manufacturer’s protocol. Transfected cells were seeded into 24-well plates (1 × 105 cells/well) and cultured in a humidified incubator containing 5% CO2 at 37°C for 24 h. The cells were subsequently resuspended in 500 µL of binding buffer containing 1% FITC-labeled Annexin V and propidium iodide. After incubation in the dark for 30 min, apoptosis levels were evaluated using the FACS Aria system (BD Immunocytometry Systems, San Jose, CA, USA) and analyzed by Cell Quest software (Becton Dickinson Ltd). All the samples were assayed three times.
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