Annexin 5 pi apoptosis kit

Apoptosis was determined with the Annexin V/PI Apoptosis Kit (MultiSciences Biotech, Hangzhou, China). Cells were harvested and centrifuged at 2,000 × g for 5 min at 4°C. Cells were resuspended in 0.5 mL of 1× annexin binding buffer at 5 × 10⁵ cells/mL. The cells were stained with Annexin V-FITC and propidium iodide (PI) for 10 min at room temperature in the dark. Signals were immediately analyzed by the flow cytometer.

To assess apoptosis, an Annexin V/PI apoptosis kit (MultiSciences Biotech, Co., Ltd., Hangzhou, China) was used, according to the manufacturer's instructions. Briefly, following incubation in 6-well plates with osteoinductive medium in the presence of various concentrations of ZA (0, 0.01, 0.1, 1, 10 or 100 µM) for 1, 4 and 7 days, the cultured MC3T3-E1 cells were gently resuspended in binding buffer and incubated for 5 min at room temperature in the dark with 5 µl Annexin V-FITC and 10 µl PI. The AnnexinV-FITC and PI-labelled cells were analyzed using a flow cytometer (FACSort; BD Biosciences, Burlington, MA, USA). Using flow cytometry, dot plots of Annexin V-FITC, on the X-axis, against PI, on the Y-axis, were used to distinguish viable cells, which are negative for PI and Annexin V-FITC, early apoptotic cells (Annexin V-positive/PI-negative) and late apoptotic or necrotic cells (AnnexinV-FITC-positive/PI-positive staining). The resultant data was analyzed using CellQuest software version 3.1 (BD Biosciences, San Jose, CA, USA).

The apoptosis induced by DOX-loaded micelles was detected and quantified by the Annexin V-PI apoptosis kit (MultiSciences Biotech Co. Ltd). Briefly, MCF-7/ADR cells were treated with free DOX·HCl, β-CD/DOX, PEG5000-CD/DOX, PELA54/DOX, and PELA54-CD/DOX (with the DOX concentration of 2 μg/mL). After 48 hours, the control (untreated) and treated cells were harvested, washed, and resuspended in 500 μL ice-cold binding buffer, and then 5 μL Annexin V/FITC solution and 1 μL PI solution were added and incubated at room temperature in the dark for 15 minutes. Then, the cell apoptosis was detected and quantified by flow cytometer (Cytomics FC 500 MCL; Beckman Coulter, Brea, CA, USA).32 (link)

After transfection, cell apoptosis was detected using a FACSCalibur flow cytometer (BD Biosciences, MA, United States) after staining using the Annexin V/PI apoptosis kit (MultiSciences, Hangzhou, China) according to the manufacturer’s instructions. For cell cycle analysis, transfected pancreatic cancer cells were harvested, washed with cold PBS, fixed in 70% ethanol at -20 °C for 24 h, and stained with a cell cycle kit (MultiSciences, Hangzhou, China).