Aria 3 flow cytometer

Manufactured by BD

Sourced in United States

The Aria III flow cytometer is an instrument designed for cell sorting and analysis. It is capable of detecting and measuring multiple parameters of individual cells or particles in a fluid sample as they pass through a laser beam. The Aria III provides high-speed cell sorting and advanced data acquisition capabilities.

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Lab products found in correlation

Polyethylenimine (pei), by Merck Group (4 mentions) Lsr 2 flow cytometer, by BD (2 mentions) Aldefluor assay kit, by STEMCELL (2 mentions) Cytoflex lx flow cytometer, by Beckman Coulter (2 mentions) Interleukin 2 (il 2), by R&D Systems (2 mentions) Fitc annexin 5 apoptosis detection kit, by BD (2 mentions) Dnase 1, by Roche (2 mentions) Live dead fixable aqua dead cell stain, by Thermo Fisher Scientific (2 mentions) Lsrii flow cytometer, by BD (2 mentions) Alexa fluor 488 annexin 5 dead cell apoptosis kit, by Thermo Fisher Scientific (2 mentions) Canto 2 flow cytometer, by BD (2 mentions) Trypsin edta, by Thermo Fisher Scientific (2 mentions) Attune flow cytometer, by Thermo Fisher Scientific (2 mentions) Tgf β, by Thermo Fisher Scientific (1 mentions) Aria flow cytometer, by BD (1 mentions) Anti cd3 fitc, by BioLegend (1 mentions) Anti ly6g pe, by BioLegend (1 mentions) Mouse tumor dissociation kit, by Miltenyi Biotec (1 mentions) Red blood cell lysis buffer, by BioLegend (1 mentions) Intracellular staining perm wash buffer, by BioLegend (1 mentions)

Spelling variants of this product

Aria III (342 citations) Aria flow cytometer (43 citations) Aria III cytometer (10 citations) BD FACS Aria II/III or Fusion flow cytometers (2 citations)

66 protocols using aria 3 flow cytometer

Characterizing NSCLC Cell Surface Markers

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NSCLC cells were trypsinized, resuspended in PBS containing 1% BSA, and stained lively with the APC-conjugated E-cadherin antibody (Miltenyi Biotec, Cat. No. 130-099-723) or APC-conjugated N-cadherin antibody (Miltenyi Biotec, Cat. No. 130-116-171) at a concentration recommended by the manufacturer. Corresponding isotype controls were used to set negative gates. Cells were either analyzed by a BD LSR II Flow cytometer, or sorted by a BD Aria III Flow Cytometer. All data were analyzed and visualized by Flowjo.
For cellular ALDH activity analysis and ALDH activity-based cell sorting, the ALDEFLUOR assay kit (STEMCELL Technologies) was used following the manufacturer's protocol with a minor modification. In brief, cells with or without drug treatment were grown to at least 70% confluence. After trypsinized, 1×10⁶/ml cells were resuspended in ALDEFLUOR buffer. Cells were then incubated with 2.5 µl ALDEFLUOR reagent per 1×10⁶ cells for 45 minutes at 37 °C. One portion of cells from each sample was treated with 5 μl ALDH inhibitor, diethylaminobenzaldehyde (DEAB, 15 μM), to set the negative gate. After incubation, cells were washed once with ALDEFLUOR assay buffer and eventually resuspended in 500 µl of ALDEFLUOR assay buffer. Samples were next subjected to a BD LSR II Flow cytometer for analysis, or a BD Aria III Flow Cytometer for sorting.

Cai S., Li N., Bai X., Liu L., Banerjee A., Lavudi K., Zhang X., Zhao J., Venere M., Duan W., Zhang J., Welliver M.X., He K, & Wang Q.E. (2022). ERK inactivation enhances stemness of NSCLC cells via promoting Slug-mediated epithelial-to-mesenchymal transition. Theranostics, 12(16), 7051-7066.

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Quantification of HbF+ and Erythroid Differentiation

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For HbF^+ve cell analysis, 1 × 10⁵ erythroid differentiated cells were briefly washed with PBS and fixed with 0.05% glutaraldehyde for 10 min and permeabilized with 0.1% Triton X-100 for 5 min. The cells were stained with anti-HbF APC antibody (dilution 1:50) and were acquired and analyzed using Cytoflex LX Flow Cytometer (Beckmann Coulter) or AriaIII flow cytometer (BD Biosciences) and analyzed using FlowJo (BD Biosciences). For erythroid differentiation analysis, 1 × 10⁵ cells from the terminal day of erythroid differentiation were stained for erythroid differentiation markers anti-CD71-FITC (dilution 1:33), anti-CD235a PE-Cy7 (dilution 1:50) and Hoechst 33342 (dilution 1:1,000). After 20 min of incubation in the dark, the cells were washed with PBS followed by analysis using Cytoflex LX Flow Cytometer (Beckmann Coulter) or AriaIII flow cytometer (BD Biosciences).

Venkatesan V., Christopher A.C., Rhiel M., Azhagiri M.K., Babu P., Walavalkar K., Saravanan B., Andrieux G., Rangaraj S., Srinivasan S., Karuppusamy K.V., Jacob A., Bagchi A., Pai A.A., Nakamura Y., Kurita R., Balasubramanian P., Pai R., Marepally S.K., Mohankumar K.M., Velayudhan S.R., Boerries M., Notani D., Cathomen T., Srivastava A, & Thangavel S. (2023). Editing the core region in HPFH deletions alters fetal and adult globin expression for treatment of β-hemoglobinopathies. Molecular Therapy. Nucleic Acids, 32, 671-688.

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Apoptosis Analysis of Lung Cancer Cells

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A549, H1975, and HCC827 cells (1 × 10⁵ cells/well) were seeded in a six-well plate for 24 h and then treated with the indicated concentrations of P.A for an additional 24 h at 37 °C. The cells were washed once with ice-cold 1× PBS and then harvested. Then, the cell pellets were re-suspended in 100 μl of 1× binding buffer. The cells were double-stained with 2 μl each of Annexin-V FITC and PI (100 μg/ml) for 15 min at room temperature (RT) in the dark. Apoptotic cells were quantified on a BD Aria III Flow Cytometer (BD Biosciences, San Jose, California, USA)

Li R.Z., Fan X.X., Duan F.G., Jiang Z.B., Pan H.D., Luo L.X., Zhou Y.L., Li Y., Yao Y.J., Yao X.J., Leung E.L, & Liu L. (2018). Proscillaridin A induces apoptosis and suppresses non-small-cell lung cancer tumor growth via calcium-induced DR4 upregulation. Cell Death & Disease, 9(6), 696.

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Isolation and Induction of Treg Precursors

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Treg precursors were defined as CD4⁺CD8⁻CD25⁺CD69⁺CD24⁺ cells that were isolated from the thymocytes of WT or RelB^−/− mice and sorted using a BD Aria III flow cytometer (BD Biosciences, CA, USA). The purity of Treg precursor cells was routinely above 90%. Cells were harvested and stimulated with IL-2 (50 U/mL; R&D) for 3 days to induce the Treg phenotype. Single-cell suspensions were collected, stained, and detected using flow cytometry.

Zhou S., Wu W., Wang Z., Wang Z., Su Q., Li X., Yu Y., Zhang W., Zhu M, & Lin W. (2020). RelB regulates the homeostatic proliferation but not the function of Tregs. BMC Immunology, 21, 37.

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In vitro Treg Induction Assay

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Naive CD4⁺ CD25⁻ T cells from WT or RelB^−/− splenocytes were sorted using a BD Aria III flow cytometer (BD Biosciences, CA, USA). The purity of CD4⁺CD25⁻ T cells was routinely above 90%, and these cells were cultured in 96-well plates with TGF-β (5 ng/mL; Peprotech) and IL-2 (50 U/mL; R&D) stimulation. Three days later, flow cytometry was used to measure the expression of CD4⁺ Foxp3⁺ T cell (iTreg).

Zhou S., Wu W., Wang Z., Wang Z., Su Q., Li X., Yu Y., Zhang W., Zhu M, & Lin W. (2020). RelB regulates the homeostatic proliferation but not the function of Tregs. BMC Immunology, 21, 37.

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Isolating Immune Cell Subsets from Murine Tissues

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Bone marrow and splenocytes were obtained from 5-month-old Vav-iCre^+/− and Vav-iCre^+/−;Ercc1^−/fl mice. Samples were incubated in ACK lysis buffer to lyse red blood cells before being washed in 1× DPBS before incubating in Fc block for 15 min on ice. Splenocytes were stained with CD3-PE and NK1.1-FITC for 30 min on ice to sort T and natural killer cells. Bone marrow was stained with CD19-APC, B220-FITC, F4/80-PE-Cy7 and CD11b-PE purchased from either BD Pharmagin or eBioscience) to sort B cells and macrophages. Cells (5 × 10⁴) were sorted into FBS using a BD Aria III flow cytometer. Sorted cells were washed in 1× DPBS and snap frozen in liquid nitrogen. Total RNA was isolated from cells using RNeasy kit and analysed for the expression of senescence and SASP markers as described above.

Yousefzadeh M.J., Flores R.R., Zhu Y., Schmiechen Z.C., Brooks R.W., Trussoni C.E., Cui Y., Angelini L., Lee K.A., McGowan S.J., Burrack A.L., Wang D., Dong Q., Lu A., Sano T., O’Kelly R.D., McGuckian C.A., Kato J.I., Bank M.P., Wade E.A., Pillai S.P., Klug J., Ladiges W.C., Burd C.E., Lewis S.E., LaRusso N.F., Vo N.V., Wang Y., Kelley E.E., Huard J., Stromnes I.M., Robbins P.D, & Niedernhofer L.J. (2021). An aged immune system drives senescence and ageing of solid organs. Nature, 594(7861), 100-105.

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ALDEFLUOR Assay for ALDH Activity

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The ALDEFLUOR Assay kit (STEMCELL Technology) was used to analyze ALDH activity in cells, and sort ALDH-dim (ALDH^dim) and ALDH-bright (ALDH^br) cells by using flow cytometry. Briefly, cells were incubated with ALDEFLUOR reagents at 37°C for 45 min according to the manufacture’s instruction. For each sample, one portion of cells was treated with 50 mM diethylaminobenzaldehyde (DEAB) to define the negative gate. After incubation, ALDEFLUOR reagents were removed; cells were re-suspended in assay buffer and subjected to a BD LSR II Flow cytometer for analysis, or a BD Aria III Flow Cytometer for sorting.

Liu L., Cai S., Han C., Banerjee A., Wu D., Cui T., Xie G., Zhang J., Zhang X., McLaughlin E., Yin M., Backes F.J., Chakravarti A., Zheng Y, & Wang Q.E. (2019). ALDH1A1 contributes to PARP inhibitor resistance via enhancing DNA repair in BRCA2−/− ovarian cancer cells. Molecular cancer therapeutics, 19(1), 199-210.

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Annexin-V FITC/PI Apoptosis Assay

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H1975 cells (1 × 10⁵ cells /well) were seeded in a 6-well plate for 24 h, and treated with the indicated concentrations of D561-0775 for an additional 48 hours at 37°C. After 48 h, the cells were washed by ice-cold 1 × PBS once and harvested by trypsinization. Then cells were centrifuged, collected and resuspended in ice-cold 1 × PBS. After removing the supernatants, cell pellets were re-suspended in 100 μl 1× Annexin -binding buffer. The cells were then double-stained with Annexin-V FITC and PI (100 μg/mL) of 2 μl respectively for 15 min at room temperature in dark. After that, 300 μl 1× Annexin-binding buffer was added to re-suspension. Apoptotic cells were quantitatively counted by a BD Aria III Flow Cytometer (BD Biosciences, San Jose, California, USA).

Chen X., Xie C., Fan X.X., Jiang Z.B., Wong V.K., Xu J.H., Yao X.J., Liu L, & Leung E.L. (2017). Novel direct AMPK activator suppresses non-small cell lung cancer through inhibition of lipid metabolism. Oncotarget, 8(56), 96089-96102.

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Annexin V-FITC Apoptosis Assay in A549 Cells

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Apoptosis was measured using an annexin V-FITC apoptosis detection kit (BD Biosciences, San Jose, CA, USA). Briefly, A549 cells (1 × 10⁵ cells/well) were cultured in a 6-well plate and transfected with control AMO or AMO-421 according to the transfection protocol. Apoptosis was detected after the addition of paclitaxel. Cells were trypsinized, washed 3 times with PBS, stained with 100 µl of binding buffer containing 2 µl of annexin V FITC (2.5 µg/ml) and 5 µl of propidine iodide (50 µg/ml) and incubated in the dark at room temperature for 15 min. The stained cells were measured quantitatively using a BD Aria III flow cytometer (BD Biosciences, San Jose, CA, USA). The data were analysed by Flow Jo software.

Duan F.G., Wang M.F., Cao Y.B., Dan L.i., Li R.Z., Fan X.X., Khan I., Lai H.L., Zhang Y.Z., Hsiao W.W., Yao X.J., Wu Q.B., Liu L., Tang Y.J, & Leung E.L. (2019). MicroRNA-421 confers paclitaxel resistance by binding to the KEAP1 3′UTR and predicts poor survival in non-small cell lung cancer. Cell Death & Disease, 10(11), 821.

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Immunofluorescent Detection of Influenza A Virus NP

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Timing: 3 h

This step describes how to monitor IAV presence in cells through the immunofluorescent detection of IAV nucleoprotein (NP).

51.

Prepare samples as described in steps 42–45c.

52.

Fix the cells by resuspending the pellet in 100 μL of 4% PFA, vortex the tubes, and incubate for 10 min at RT.

53.

Wash the cells with 1 mL of FACS buffer.

54.

Permeabilize the cells by resuspending the pellet in 100 μL of 0.5% saponin.

55.

Minimize intracellular unspecific antibody binding by incubation with an anti-mouse CD16/32 antibody (Fc blocking diluted 1:100) for 15 min at 4°C.

56.

Stain the cells intracellularly by the addition of FITC-conjugated influenza A anti-NP antibody (10 μg/mL) directly in the cell suspension for 30 min at 4°C in the dark.

57.

At the end of the incubation, wash the cells once with permeabilization buffer and then with FACS buffer.

58.

Analyze on a BD ARIA III flow cytometer or equivalent. Unstained samples and single-stained samples are used to set appropriate PMT voltages for the positive and negative populations.

Galani I.E., Triantafyllia V., Eleminiadou E.E, & Andreakos E. (2022). Protocol for influenza A virus infection of mice and viral load determination. STAR Protocols, 3(1), 101151.

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