Apc labeled annexin 5

Manufactured by BD

Sourced in United States

APC-labeled Annexin V is a fluorescently-labeled protein that binds to phosphatidylserine, a cellular component exposed during apoptosis. It can be used to detect and quantify apoptotic cells.

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Lab products found in correlation

7 aad, by BD (2 mentions) Facscalibur, by BD (2 mentions) Recombinate, by Baxter (1 mentions) Propidium iodide, by BD (1 mentions) Flow cytometry, by BD (1 mentions) Fitc labeled anti cd56, by BD (1 mentions) Fitc labeled anti cd25, by BD (1 mentions) Apc labeled anti cd45ro, by BD (1 mentions) Facsaria flow cytometer, by BD (1 mentions) Pkh67 green fluorescent cell linker dye, by Merck Group (1 mentions) Wt1126 134 peptide, by JPT Peptide Technologies (1 mentions) Aim 5 medium, by Thermo Fisher Scientific (1 mentions) Facsaria 2, by BD (1 mentions)

8 protocols using apc labeled annexin 5

Evaluating FVIII and ADAMTS-13 Interactions

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Full length FVIII was either a kind gift from CSL-Behring (Helixate® NexGen, Marburg, Germany) or from Baxter (Recombinate®, Maurepas, France). Recombinant human A disintegrin and metalloprotease with thrombospondin type I repeats-13 (ADAMTS-13) was a kind gift from Baxter. Complement human proteins Factor B, Factor D, C3, C3b and C3-depleted serum were purchased from Complement Technology (Comptech, TX, USA) and Merck Millipore (Merck Chemicals Ltd., Nottingham, UK). Human serum was obtained from AB blood type healthy donors. Antibodies against CD1a, CD3, CD14, CD40, CD83, CD86, HLA-DR, CD206, low density lipoprotein receptor-related protein (LRP, CD91), CD209, CD68 and APC-labeled Annexin V were purchased from BD Pharmingen (San Jose, CA, USA). Antibody against CD20 was purchased from eBiosciences (San Diego, CA, USA).
The biotinylated monoclonal mouse anti-human FVIII antibody GMA-8015 and sheep polyclonal anti-human FVIII (SAF8C) were from Green Mountain Antibodies (Burlington, VT, USA) and Affinity Biological (Ancaster, Canada). The monoclonal anti-human FVIII antibody 77IP52H7 was a kind gift from LFB (Les Ulis, France). The biotinylated monoclonal mouse anti-human ADAMTS-13 antibody (20A5) and polyclonal goat anti-mouse C3b/iC3b (clone A209) were from Clinisciences (Nanterre, France) and Quidel (San Diego, USA), respectively.

Rayes J., Ing M., Delignat S., Peyron I., Gilardin L., Vogel C.W., Fritzinger D.C., Frémeaux-Bacchi V., Kaveri S.V., Roumenina L.T, & Lacroix-Desmazes S. (2018). Complement C3 is a novel modulator of the anti-factor VIII immune response. Haematologica, 103(2), 351-360.

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Apoptosis Induction Assay with THZ-P1-2

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In total, 1 × 10⁵ cells per well were seeded in a 24-well plate in an FBS-supplemented medium in the presence of vehicle or THZ-P1-2 (1.6, 3.2, and 6.4 μM) for 24 h. Next, the cells were washed with ice-cold phosphate-buffered saline (PBS) and resuspended in a binding buffer containing 1 μg/mL propidium iodide (PI) and 1 μg/mL APC-labeled annexin V (BD Pharmingen, San Diego, CA, USA). All specimens were analyzed using flow cytometry (FACSCalibur) after incubation for 15 min at room temperature in a light-protected area. Ten thousand events were recorded for each sample.

Lima K., Pereira-Martins D.A., de Miranda L.B., Coelho-Silva J.L., Leandro G.D., Weinhäuser I., Cavaglieri R.D., Leal A.D., da Silva W.F., Lange A.P., Velloso E.D., Griessinger E., Hilberink J.R., Ammatuna E., Huls G., Schuringa J.J., Rego E.M, & Machado-Neto J.A. (2022). The PIP4K2 inhibitor THZ-P1-2 exhibits antileukemia activity by disruption of mitochondrial homeostasis and autophagy. Blood Cancer Journal, 12(11), 151.

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Annexin V-PI Apoptosis Assay

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A total of 1 × 10⁵ cells per well were seeded in 24-well plates in the presence of a vehicle or eribulin (0.25, 0.5, and 1 nM) for 72 h. The cells were then washed twice with ice-cold PBS and resuspended in a binding buffer containing 1 μg/mL propidium iodide (PI), and 1 μg/mL APC-labeled annexin V (BD Biosciences, San Jose, CA, USA). All of the specimens were acquired by flow cytometry (FACSCalibur; Becton Dickinson, Franklin Lakes, NJ, USA) after incubation for 15 min at room temperature in a light-protected area, and they were analyzed using FlowJo software (Treestar, Inc., San Carlos, CA, USA).

Vicari H.P., Lima K., Costa-Lotufo L.V, & Machado-Neto J.A. (2022). Cellular and Molecular Effects of Eribulin in Preclinical Models of Hematologic Neoplasms. Cancers, 14(24), 6080.

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Piperine-Induced Apoptosis in DU145 Cells

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DU145 cells were planted at a concentration of 1×10⁶/ml onto 6-well culture plates. While the cells achieved about 70% degree confluence, the medium was exchanged, and piperine (160 µM) or diluent was supplemented. Subsequently, DU145 cells were trypsinized to obtain a single cell suspension. One million cells were double-stained with APC-labeled annexin V and propidium iodide (BD Biosciences, Franklin Lakes, NJ, USA). Proportion of apoptotic cells was determined by flow cytometry (BD Biosciences).

Zeng Y, & Yang Y. (2018). Piperine depresses the migration progression via downregulating the Akt/mTOR/MMP-9 signaling pathway in DU145 cells. Molecular Medicine Reports, 17(5), 6363-6370.

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Quantifying AT1-AA-producing B cells

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CD19⁺CD5⁺ B cells in the peripheral blood of mice were stained with APC-conjugated Rat Anti-Mouse CD19 and PE-conjugated Rat Anti-Mouse CD5, and subjected to flow cytometry analysis (BD FACS Calibur) to examine the percentage of AT1-AA-producing CD19⁺CD5⁺ B cells. Cultured trophoblast cells were stained with 7-AAD and APC-labeled Annexin V (BD Pharmingen) according to the manufacturer’s instructions, and subjected to flow cytometry analysis (BD FACS Calibur) to examine trophoblast cell apoptosis. The data were analyzed by FlowJo software.

Liu H., Cheng F., Xu Q., Huang W., Wang S., Sun R., Ye D, & Zhang D. (2020). Lipoxin A4 suppresses angiotensin II type 1 receptor autoantibody in preeclampsia via modulating caspase-1. Cell Death & Disease, 11(1), 78.

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Early Apoptosis Detection by Flow Cytometry

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Flow cytometry with 7-AAD and APC-labeled Annexin V (BD Pharmingen, USA) was executed in line with the protocol to recognize phosphatidylserine externalization, a sign of early apoptosis.

Yun W.J., Xue H., Yang N., Xiao L.J., Sun H.Z, & Zheng H.C. (2023). Oncogenic roles of GPR176 in breast cancer: a potential marker of aggressiveness and a potential target of gene therapy. Clinical & Translational Oncology, 25(10), 3042-3056.

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Characterization of Activated T Cells

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After 7 days of expansion, T cells were harvested and immunophenotyping was performed. T cells were incubated with the following monoclonal antibodies: fluorescein isothiocyanate (FITC)-, phycoerythrin (PE)-, PE-cyanine 7 (PE-Cy7)-, and V450-labeled anti-CD3; allophycocyanin (APC)-, and Alexa Fluor 700-labeled anti-CD4; Peridinin-chloropyll-protein complex (PerCP)-labeled anti-7-Aminoactinomycin D (7-AAD); FITC-labeled anti-CD28; FITC-labeled anti-CD69; FITC-labeled anti-CD94; FITC-labeled anti-CD56; FITC-labeled anti-TCRαβ; FITC-labeled anti-CD95; APC-labeled anti-CD45RO; PE-Cy7-labeled anti-CCR7; FITC-labeled anti-CD25; APC-labeled Annexin V (all from BD Bioscience, Franklin Lakes, NJ, USA); FITC-labeled anti-TCRγδ (Becman Coulter, Fullerton, CA, USA); APC-Alexa Fluor 700-labeled anti-CD127 (Beckman Coulter, Fullerton, CA, USA); Pacific Orange-labeled anti-CD8 (Invitrogen); PE-labeled anti-CD39 (BD Bioscience, Franklin lakes, NJ, USA) for 15 min and washed before flow cytometry (FACS Aria flow cytometer, BD Biosciences) and analysis (FlowJo software, Tree Star, Inc., Ashland, OR, USA). T cell death was assessed by flow cytometry staining for 7-AAD and Annexin V after 7 days culture with beads and cytokines followed by a brief (6 h) period of stimulation with peptide mix spanning the cytomegalovirus (CMV) protein pp65.

Tauriainen J., Gustafsson K., Göthlin M., Gertow J., Buggert M., Frisk T.W., Karlsson A.C., Uhlin M, & Önfelt B. (2015). Single-Cell Characterization of in vitro Migration and Interaction Dynamics of T Cells Expanded with IL-2 and IL-7. Frontiers in Immunology, 6, 196.

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Cytotoxicity Assay for CD8+ T cells

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The killing capacity of electroporated human primary resting CD8⁺ T cells against T2 cells was determined using a flow cytometry-based protocol as described previously with minor modifications (51 (link)). Briefly, prior to co-culture tumor cells were stained with PKH67 green fluorescent cell linker dye (Sigma-Aldrich) according to the manufacturer's protocol. PKH67⁺ T2 cells were incubated with WT1₃₇₋₄₅ or WT1_126−134 peptide (JPT Peptide Technologies) in AIM-V medium (Gibco Invitrogen) for 90 min at room temperature under constant motion. Next, T2 cells were cultured alone or with electroporated human primary resting CD8⁺ T cells for 6 h at an effector-target ratio of 20:1. After co-culture, samples were stained with propidium iodide (PI) and APC-labeled annexin V (BD Biosciences). Samples were analyzed using a FACSAria II flow cytometer (BD Biosciences). Cytotoxicity was calculated based on the survival of PKH67⁺ T2 cells using the following equation:

Campillo-Davo D., Fujiki F., Van den Bergh J.M., De Reu H., Smits E.L., Goossens H., Sugiyama H., Lion E., Berneman Z.N, & Van Tendeloo V. (2018). Efficient and Non-genotoxic RNA-Based Engineering of Human T Cells Using Tumor-Specific T Cell Receptors With Minimal TCR Mispairing. Frontiers in Immunology, 9, 2503.

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