CMLT1 as CD26+ cells and HL60 as CD26− cells were treated with free venetoclax and IL-VX. 1 × 104 cells were plated in 96 wells plate and treated with different concentrations of the free venetoclax (ranging from 10 nM to 1 µM) and IL-VX (ranging from 10 nM to 1 µM). After 48 h, an apoptosis assay by Annexin-V kit (Miltenyi Biotech, Bergisch Gladbach, Germany) was performed according to manufacturer protocol. Briefly, cells were collected and washed with Annexin buffer and after addition of Annexin-V FITC were incubated at RT for 15 min in the dark. Afterward, cells were washed again with Annexin buffer and Propidium Iodide (PI) was added. Apoptosis assays were acquired by FACSVERSE (BD-Bioscience) and were analyzed by Kaluza software version 2.1 (Beckman Coulter Fullerton, Brea, CA, USA).
Annexin 5 kit
Manufactured by Miltenyi Biotec
Sourced in Germany
The Annexin-V kit is a laboratory reagent used for the detection and quantification of apoptosis, or programmed cell death. The kit contains Annexin-V, a protein that binds to phosphatidylserine, a molecule expressed on the surface of apoptotic cells. This binding can be detected and measured, providing a means to assess the extent of apoptosis in a sample.
Automatically generated - may contain errors
Lab products found in correlation
Kaluza software version 2, by Beckman Coulter (1 mentions)
Facsverse, by BD (1 mentions)
Macsquant analyzer, by Miltenyi Biotec (1 mentions)
Trypan blue, by Merck Group (1 mentions)
Cellquest software, by BD (1 mentions)
Facscalibur, by BD (1 mentions)
Cfse cell proliferation kit, by Thermo Fisher Scientific (1 mentions)
4 protocols using annexin 5 kit
1
Apoptosis Induction by Venetoclax and IL-VX
2
T Cell Apoptosis in HSC Co-culture
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
CD34+ HSCs and MNCs were prepared as described above. T cells (1×106) were cultured alone or co-cultured with CD34+ HSCs (1×105) in ratio of 1:10 or stimulated with PHA (50 mg/ml final concentration) for 3 days, harvested and quantified, stained with Annexin V kit (Miltenyi Biotec, Germany) and analyzed by flow cytometry (MACSQuant Analyzer, 2440).
3
Cell Proliferation, Apoptosis, and Cell Cycle Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
For proliferation assay 2 × 105 cells were seeded in a 12-well plate. The count of viable cells was performed after 72 and 120 h using trypan blue (Sigma-Aldrich, City of Saint Louis, MO, USA). Each experiment was performed in triplicate, considering as standard output the mean value ± standard deviation.
Apoptosis was quantified by the percentage of Annexin Vpos/7AADneg cells using flow-cytometry. Following the manufacturer’s instructions, the Annexin V Kit (Miltenyi Biotec, Bergisch Gladbach, Germany) was adopted to evaluate the percentage of apoptotic cells 48 h after miRNA transfection.
Cell cycle was evaluated 48 h after miRNA transfection. After fixation with 70% cold ethanol, cells were stained with propidium iodide (50 µg/mL) for 40 min. Analyses were carried out by flow-cytometry using BD FACS Calibur and Cell Quest software (BD Biosciences, Franklin Lakes, NJ, USA).
Apoptosis was quantified by the percentage of Annexin Vpos/7AADneg cells using flow-cytometry. Following the manufacturer’s instructions, the Annexin V Kit (Miltenyi Biotec, Bergisch Gladbach, Germany) was adopted to evaluate the percentage of apoptotic cells 48 h after miRNA transfection.
Cell cycle was evaluated 48 h after miRNA transfection. After fixation with 70% cold ethanol, cells were stained with propidium iodide (50 µg/mL) for 40 min. Analyses were carried out by flow-cytometry using BD FACS Calibur and Cell Quest software (BD Biosciences, Franklin Lakes, NJ, USA).
4
CD8+ T Cell Proliferation and Apoptosis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
For determination of the cell proliferation, CD8+ T cells were stained with 10 µM CellTrace™ Carboxyfluorescein succinimidyl ester (CFSE) Cell Proliferation Kit (Thermo Fisher) as described [50 (link)], then seeded at a density of 106 cells/ml and cultured for five days. After this incubation, CD8+ T cells were washed once with PBS and directly analyzed by flow cytometry. The flow cytometric data were processed using FCS Express 6 Plus Research Edition (San Diego, CA, USA) with the software’s standard specification for proliferation assays. Results are presented as proliferation indexes indicating the average number of times the cells have divided.
For the determination of the apoptosis rate, CD8+ T cells either left untreated or treated for the indicated time were stained with an annexin-V kit (Miltenyi) according to the manufacturers’ protocol followed by flow cytometric analysis. Staining with a FITC labelled anti-annexin-V mAb and propidium iodide (PI) was used to discriminate between alive cells (annexin−, PI−), debris (annexin− PI+), dead cells (annexin+ PI+) and apoptotic cells (annexin+ PI−).
For the determination of the apoptosis rate, CD8+ T cells either left untreated or treated for the indicated time were stained with an annexin-V kit (Miltenyi) according to the manufacturers’ protocol followed by flow cytometric analysis. Staining with a FITC labelled anti-annexin-V mAb and propidium iodide (PI) was used to discriminate between alive cells (annexin−, PI−), debris (annexin− PI+), dead cells (annexin+ PI+) and apoptotic cells (annexin+ PI−).
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!