CPB cells at six-well plates were infected with 1 MOI of the RGNNV isolated from diseased red-spotted grouper or equal volume of PBS as negative control. At 24 h p.i., cells were used for apoptosis assay using annexin V-FITC/PI apoptosis detection kit (TransGen Biotech, Beijing, China) as described previously [24 (link)]. For DAPI staining, cells were treated as described before [24 (link)]. Finally the cells were viewed using an inverted fluorescence microscope.
Annexin 5 fitc pi apoptosis detection kit
Manufactured by Transgene
Sourced in China, United States
The Annexin V-FITC/PI apoptosis detection kit is a laboratory equipment product that is used to detect and quantify apoptosis, or programmed cell death, in cell samples. The kit contains Annexin V-FITC and propidium iodide (PI) as the key detection reagents. Annexin V-FITC binds to phosphatidylserine, which is exposed on the cell surface during apoptosis, while PI stains the nucleus of cells with compromised cell membranes, indicating late apoptosis or necrosis. This kit allows for the identification and quantification of early and late apoptotic cells using flow cytometry or fluorescence microscopy.
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Lab products found in correlation
L glutamine, by Thermo Fisher Scientific (2 mentions)
Erlotinib hcl osi 744, by Selleck Chemicals (1 mentions)
Cell cycle analysis kit, by Keygen Biotech (1 mentions)
Modfit software, by Verity Software House (1 mentions)
Mitotracker, by Thermo Fisher Scientific (1 mentions)
Hif 1α 36169, by Cell Signaling Technology (1 mentions)
Mitosox, by Thermo Fisher Scientific (1 mentions)
Cck 8 assay kit, by Meilun (1 mentions)
Facscan, by BD (1 mentions)
Thapsigargin, by Beyotime (1 mentions)
Bortezomib ps 341, by Selleck Chemicals (1 mentions)
Lipofectamine 3000, by Thermo Fisher Scientific (1 mentions)
Fluo 4 am, by Beyotime (1 mentions)
Fura 2 am, by Beyotime (1 mentions)
7 protocols using annexin 5 fitc pi apoptosis detection kit
1
Apoptosis Induction in CPB Cells by RGNNV
2
Apoptosis and Cell Cycle Analysis
3
Annexin V-FITC/PI Apoptosis Detection
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For flow cytometry (FCM) detection, an Annexin V-FITC/PI Apoptosis Detection Kit (TransGen Biotech, Beijing, China) was used to detect the early apoptosis levels. The experiment was performed according to the manufacturer’s protocol. The negative control (without Annexin V and PI) and positive controls (with Annexin V or PI) were also designed according to the protocol. After the examples were prepared, the fluorescence signal was detected within 1 h by FCM (BD). Annexin V-FITC was detected at a 488-nm excitation wavelength and a 530-emission wavelength. Propidium iodide was detected at 488 nm excitation wavelength and a 630-nm emission wavelength.
4
Annexin V Apoptosis Assay Protocol
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Collect each group of test cells, wash the cells with pre-cooled PBS, add 300 μl of binding buffer to resuspend the cells, and adjust the cell density to 1 × 106 cells/ml. According to the manufacturer’s recommendations, Annexin V-FITC/PI Apoptosis Detection Kit (TransGen Biotech) was used for apoptosis analysis. Take 100ul of cell suspension into the flow tube, and add 5ul each of Annexin V-FITC and PI. After mixing, incubate at room temperature for 15 min in the dark, and add 400 ul of PBS to the reaction tube. Ensure that the cell apoptosis detected by flow cytometry within 1 h. The ModFit software (Verity Software House) used to analyze flow cytometry data.
5
Comprehensive Cellular Analysis Protocol
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The antibodies to CISD2 (60758 S), IRP2 (37135S), and HIF-1α (#36169) were obtained from Cell Signaling Technology (Danvers, MA, USA). The antibodies to Transferrin Receptor (sc-32272) were obtained from Santa Cruz (Dallas, TX, USA). The antibodies to β-Actin (ab8226) were obtained from Abcam (Cambridge, MA, USA). CCK-8 Assay Kit was obtained from Meilunbio (Dalian, China). C11-BODIPY (581/591), deferoxamine (DFO), Rhodamine B-[(1, 10-phenanthroline-5-yl)-aminocarbonyl] benzyl ester (RPA), DCF-DA, MitoTracker, and MitoSOX were obtained from Thermo Fisher Scientific (Waltham, MA, USA). The BeyoClick™ EdU Cell Proliferation Kit and ATP detection kit purchased from Beyotime (Shanghai, China), Annexin V-FITC/PI apoptosis detection kit was purchased from TransGen Biotech (Beijing, China).
6
Annexin V-FITC/PI Apoptosis Assay
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Cell apoptosis was measured through the Annexin V-FITC/PI Apoptosis Detection Kit (TransGen, Perking, China). Briefly, cells were washed and collected 48 h after transfection. Next, Annexin V-FITC/PI was used to double stain the cells. Subsequently, flow cytometry (FACScan; BD Biosciences, Franklin Lakes, NJ, USA) was manipulated to monitor the apoptosis.
7
Apoptosis Induction and Modulation Assay
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DMEM, RPMI-1640 medium, and fetal bovine serum (FBS) were purchased from HyClone, L-glutamine from Gibco, Propidium Iodide (PI) Solution (421301) from Biolegend (CA, USA), Annexin V-FITC/PI apoptosis detection kit (FA101-02) from TransGen Biotech (Beijing, China), Lipofectamine 3000 (L3000015) from Invitrogen (CA, USA), Thapsigargin (CHXSC0389), MG-132 (S1748), Fura-2 AM (S1052) and Fluo-4 AM (S1060) from Beyotime (Jiangsu, China), Chloroquine (CQ, 105M4035V) from Sigma, Bortezomib (PS-341, S1013), BafilomycinA1 (BAF, S1413) from Selleckchem (PA, USA), Cycloheximide (CHX, C112766) from Aladdin (Shanghai, China).
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