The induced M2 macrophages were probed with F4/80 (1:500), CD86 (1:500), CD206 (1:200), or isotype immunoglobulin G 2b (1:300, all from Abcam). Then, the M2 macrophages (1 × 106 cells/mL) were detected on a flow cytometer (FACSCalibur, BD Biosciences) for measuring the proportion of CD86+ and CD206+ cells. M2 macrophages were F4/80+ and CD206+. Cell apoptosis was tested by Annexin V-fluorescein isothiocyanate (FITC)/propidium iodide (PI) apoptosis detection kit (Invitrogen). PANC-1 and AsPC-1 cells, and P-CSCs and A-CSCs were added with dimethyl sulfoxide (10 μmol/mL) or anti-tumor drug perifosine (10 μmol/mL) (Zhou et al. 2020 (link)). Treated for 48 h, cells were fixed in 70% ethanol, adjusted to 1 × 106 cells/mL with 1× Binding Buffer, and stained by 5 μL Annexin V-FITC and 1 μL PI (Invitrogen). The flow cytometer (FACSCalibur) was adopted to apoptosis detection.
Annexin 5 fluorescein isothiocyanate fitc propidium iodide apoptosis detection kit
Manufactured by Thermo Fisher Scientific
Sourced in United States
The Annexin V-FITC/PI apoptosis detection kit is a laboratory tool that enables the identification and quantification of apoptotic cells. This kit utilizes the binding properties of Annexin V, a calcium-dependent phospholipid-binding protein, which has a high affinity for the phospholipid phosphatidylserine. The kit also includes propidium iodide (PI), a DNA-binding dye. Together, these components can distinguish between viable, early apoptotic, and late apoptotic/necrotic cells through flow cytometry or fluorescence microscopy analysis.
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Lab products found in correlation
Annexin 5 fitc, by Thermo Fisher Scientific (1 mentions)
F4 80, by Abcam (1 mentions)
Cd206, by BD (1 mentions)
Cd206, by Abcam (1 mentions)
Facscalibur, by BD (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Dojindo Laboratories (1 mentions)
Live dead viability cytotoxicity kit, by Thermo Fisher Scientific (1 mentions)
2 7 dichlorodihydrofluorescein diacetate dcfh da, by Merck Group (1 mentions)
9 10 anthracenediyl bis methylene dimalonic acid abda, by Merck Group (1 mentions)
Cell counting kit 8 cck 8, by Dojindo Laboratories (1 mentions)
Cycletest plus dna reagent kit, by BD (1 mentions)
Facscan, by BD (1 mentions)
Binding buffer, by BD (1 mentions)
Facstation software, by BD (1 mentions)
Binding buffer, by Keygen Biotech (1 mentions)
Facscalibur flow cytometer, by BD (1 mentions)
4 protocols using annexin 5 fluorescein isothiocyanate fitc propidium iodide apoptosis detection kit
1
Characterization of Tumor-associated Macrophages
2
Synthesis and Characterization of Photosensitive Polymers
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The conjugated polymers poly[(9,9'-dioctyl-2,7-divinylene-fluorenylene)-alt-2-methoxy-5-(2-ethyl-hexyloxy)-1,4-phenylene] (PFPV) and poly[(9,9-dioctylfluorenyl-2,7-diyl)-co-(1,4-benzo-(2,1′,3)-thiadazole)] (PFBT) were purchased from Luminescence Technology Corp. and ADS Dyes, Inc., respectively. Tetraphenylporphyrin (TPP) was purchased from Adamas-beta, and bis[3,4,6-trichloro-2-(pentyloxycarbonyl) phenyl] oxalate (CPPO) was supplied by TCI. Poly(ethylene glycol)-poly(ε-caprolactone) (PEG-PCL) was synthesized according to previously reported methods.1 Cholesterol-PEG-folate (Mw = 2000) was purchased from Xi'an Ruixi Biological Technology Co., Ltd. 9,10-Anthracenediyl-bis(methylene) dimalonic acid (ABDA) and 2,7-dichlorodihydrofluorescein diacetate (DCFH-DA) were purchased from Sigma-Aldrich. 4',6-Diamidino-2-phenylindole (DAPI) and Cell Counting Kit-8 (CCK-8) were obtained from Dojindo Molecular Technologies. The LIVE/DEAD Viability/Cytotoxicity Kit and Annexin V-fluorescein isothiocyanate (FITC)/propidium iodide (PI) apoptosis detection kit were provided by Invitrogen. All other chemicals, if not specified, were used as received without further purification.
3
Cell Cycle and Apoptosis Analysis
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After synchronized by serum-free starvation for 24 hours and infected with adenovirus for 24h hours, HeLa cells were then harvested and stained with propidium iodide using a Cycle TEST PLUS DNA Reagent Kit (Becton Dickinson, USA). Cell cycles were analyzed using flow cytometry with a FACScan (Becton Dickinson, USA).
Cell apoptosis was detected using an Annexin V-fluorescein isothiocyanate (FITC)/propidium iodide (PI) apoptosis detection kit (Invitrogen, USA) following the manufacturer's protocol. Briefly, cells were collected, centrifuged, and re-suspended in 500μl of 1×binding buffer in tubes. After added with Annexin V-FITC and PI, tubes were incubated darkly at room temperature for 15 minutes. Cell apoptosis assay was performed immediately on flow cytometry. Each experiment was performed at least three times.
Cell apoptosis was detected using an Annexin V-fluorescein isothiocyanate (FITC)/propidium iodide (PI) apoptosis detection kit (Invitrogen, USA) following the manufacturer's protocol. Briefly, cells were collected, centrifuged, and re-suspended in 500μl of 1×binding buffer in tubes. After added with Annexin V-FITC and PI, tubes were incubated darkly at room temperature for 15 minutes. Cell apoptosis assay was performed immediately on flow cytometry. Each experiment was performed at least three times.
4
Apoptosis Detection in HepG2 Cells
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Cell apoptosis was detected with an Annexin V-fluorescein isothiocyanate (FITC)/propidium iodide (PI) apoptosis detection kit (Invitrogen; Thermo Fisher Scientific, Inc.). pcDNA3.1-pAFP-TK was transfected into HepG2 cells. Cells in the control group received no transfection and no further treatments. Following 48 h incubation, the pcDNA3.1-pAFP-TK group was treated with 150 µg/ml GCV for 2 days. Cells were harvested with 0.25% trypsin then sedimented by centrifugation at 335.4 × g for 3 min at room temperature. The supernatant was then discarded, cells were washed twice with PBS, then 100 µl binding buffer (Nanjing KeyGen Biotech Co., Ltd., Nanjing, China) was added to resuspend the cells. Cell suspension (100 µl) was then added to the flow tube, along with 5 µl Annexin V-FITC at a final concentration of 1 µg/ml and 10 µl (250 ng) of PI. The cells were mixed and incubated for 15 min at room temperature in the dark. Finally, 400 µl binding buffer was added and flow cytometry was used to detect cell apoptosis, using the BD FACSCalibur™ flow cytometer and the BD FACStation™ software (BD Biosciences, Franklin Lakes, NJ, USA).
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