Biotin conjugated annexin 5

Manufactured by BD

Biotin-conjugated Annexin V is a protein that binds to phosphatidylserine, a molecule expressed on the surface of apoptotic cells. This binding can be used to detect and quantify apoptosis in various cell-based assays.

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Lab products found in correlation

Pe conjugated streptavidin, by BD (1 mentions) Streptavidin allophycocyanin apc, by BD (1 mentions) 7aad dye, by BD (1 mentions) Facscelesta cell analyzer, by BD (1 mentions) Annexin 5 binding buffer, by BD (1 mentions) Streptavidin efluor 450, by Thermo Fisher Scientific (1 mentions) 7 aad, by Merck Group (1 mentions) Wst 1 assay, by Roche (1 mentions)

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Annexin V (870 citations)

3 protocols using biotin conjugated annexin 5

Quantification of Neutrophil Apoptosis

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The apoptotic index of neutrophils was quantified by staining the white blood cells with biotin-conjugated Annexin V (BD Bioscience). After the incubation with the saturated concentration of biotin-conjugated Annexin V, the cell suspension was further incubated with PE-conjugated Streptavidin (BD Bioscience). The positively stained apoptotic cells were counted, and the apoptotic index was calculated as the percentage of apoptotic cells within the total number of cells.

Manukyan G., Aminov R., Hakobyan G, & Davtyan T. (2015). Accelerated Apoptosis of Neutrophils in Familial Mediterranean Fever. Frontiers in Immunology, 6, 239.

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Annexin V Apoptosis Assay for A549 and Capan-I Cells

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Wild-type or MET^−/− A549 (5 × 10⁴ cells/well) or Capan-I (10⁵ cells/well) cells were seeded in six-well ultralow attachment surface plates in medium plus 2% FCS (A549) or 20% FCS (Capan-I), with or without HGF (25 ng/mL). After 24 h (A549) or 48 h (Capan-I), cell suspensions were collected, washed in PBS, and resuspended in 100 μL of Annexin V binding buffer (BD Biosciences) containing 2.5 μL of biotin-conjugated Annexin V (BD Biosciences) and incubated for 15 min at room temperature. Following one wash in PBS, cells were incubated with 0.5 μL of streptavidin–allophycocyanin (APC; BD Biosciences) in 100 μL of Annexin V binding buffer (BD Biosciences) for an additional 15 min at room temperature in the dark. Then, 7-AAD dye (BD Biosciences) was added right before the acquisition with FACSDiva v9.2 software on a BD FACSCelesta Cell Analyzer. Post-acquisition analysis was performed using FlowJo v10.8.1 software.

Modica C., Cortese M., Bersani F., Lombardi A.M., Napoli F., Righi L., Taulli R., Basilico C, & Vigna E. (2023). Genetic Ablation of the MET Oncogene Defines a Crucial Role of the HGF/MET Axis in Cell-Autonomous Functions Driving Tumor Dissemination. Cancers, 15(10), 2742.

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Cell Viability and Apoptosis Assessment

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Cell viability was assessed by a WST-1 assay (Roche) according to the manufacturer’s instructions. Briefly, 5000 cells were seeded in 96-well plates and treated with drugs as described above. After 72h, 10 μl of WST-1 reagent was added to each well and absorbance at 450 nm was measured using a TECAN plate-reader (TECAN).
Cell apoptosis was assessed by FACS analysis of biotin-conjugated AnnexinV (BD Biosciences) and 7AAD (Sigma), according to the manufacturer’s instructions. Briefly, 200000 cells were seeded in 6-well plates and treated with drugs. After 48h, the cells were trypsinized and transferred to a tube along with the conditioned medium. Cells were centrifuged at 2000 rpm, washed with ice-cold PBS, and resuspended in 1x binding buffer at a concentration of 10⁶ cells/ml. The cell suspension was then stained with biotin-conjugated AnnexinV for 15 min at room temperature, followed by streptavidin-eFluor 450 (eBioscience; 1:100) for 15 min on ice. After washing, cells were resuspended in 7AAD (1:200) and immediately analysed by FACS.

Keklikoglou I., Cianciaruso C., Güç E., Squadrito M.L., Spring L.M., Tazzyman S., Lambein L., Poissonnier A., Ferraro G.B., Baer C., Cassará A., Guichard A., Iruela-Arispe M.L., Lewis C.E., Coussens L.M., Bardia A., Jain R.K., Pollard J.W, & De Palma M. (2018). Chemotherapy elicits pro-metastatic extracellular vesicles in breast cancer models. Nature cell biology, 21(2), 190-202.

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