Facsymphony a5 laser analyzer

Manufactured by BD

Sourced in United States

The BD FACSymphony A5-Laser Analyzer is a flow cytometer designed for analyzing and sorting cells. It features five lasers and can detect up to 20 parameters simultaneously. The instrument is capable of performing high-speed cell analysis and sorting.

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Lab products found in correlation

Hanks balanced salt solution (hbss), by Thermo Fisher Scientific (1 mentions) Apc conjugated mouse anti cd44, by BD (1 mentions) Pe conjugated mouse anti cd24, by BD (1 mentions) Efluor 506, by Thermo Fisher Scientific (1 mentions) 123count ebeads, by Thermo Fisher Scientific (1 mentions) Pharm lyse buffer, by BD (1 mentions) Rat anti mouse cd11b apc cy7, by BioLegend (1 mentions) Rat anti mouse cd11c pe cy7, by BD (1 mentions) Fitc rat anti mouse cd45, by BioLegend (1 mentions) Rat anti mouse cd45 apc, by BioLegend (1 mentions) Fcr blocking reagent, by Miltenyi Biotec (1 mentions) Fixable viability dye, by Thermo Fisher Scientific (1 mentions) Cd16 32 antibody, by BioLegend (1 mentions) Lsrfortessa x 20 analyzer, by BD (1 mentions) Foxp3 transcription factor staining buffer set, by Thermo Fisher Scientific (1 mentions)

Close spelling variants of this product

BD FACSymphony A5 analyzer BD FACSymphony A5 BD FACSymphony analyzer FACSymphony

6 protocols using facsymphony a5 laser analyzer

Retinal Immune Cell Profiling by Flow Cytometry

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Eyes were enucleated and stored briefly in HBSS (14,025,076, Gibco, Carlsbad, CA). Eyes were dissected on a microscope to isolate retina alone. The retina was cut into at least four pieces and digested in Liberase TL for one hour at 37⁰C with shaking (200 RPM). Post digestion, retinal cells were treated identically to our previously published flow cytometry procedure [23 ]. Briefly, a single cell suspension was obtained and stained for innate immune cell markers (see Table 2). Samples were run on a FACSymphony A5-Laser Analyzer (Becton Dickinson, CA, USA). Experiments were performed on two separate days and combined.Table 2

Flow cytometry antibodies

Antibody	Fluorophore	Manufacturer, product number
Fc block	–	BD Biosciences, 553,142
Aqua Live/Dead	AmCyan	ThermoFisher, 65–0866-14
CD45	BUV395	BD Biosciences, 564,279
CD64	PE	BD Biosciences, 139,304
CD11b	APC-Cy7	BD Biosciences, 557,657
Ly6G	PE-CF594	BD Biosciences, 562,700
B220	PE-CF594	BD Biosciences, 562,313
NK1.1	PE-CF594	BD Biosciences, 562,864
SiglecF	PE-CF594	BD Biosciences, 562,757
CD4	PE-CF594	BD Biosciences, 562,314
CD8	PE-CF594	BD Biosciences, 562,315
Ly6C	FITC	BD Biosciences, 561,085
Cx3Cr1	Alexa Fluor 647	Biolegend, 149,004
CD206	PE-Cy7	Biolegend, 141,720

Rajesh A., Droho S, & Lavine J.A. (2022). Macrophages in close proximity to the vitreoretinal interface are potential biomarkers of inflammation during retinal vascular disease. Journal of Neuroinflammation, 19, 203.

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Isolation and Analysis of Retinal Immune Cells

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Eyes were enucleated directly into HBSS. Eyes were dissected to isolate retina and choroid/RPE/sclera complex. Retina and choroid/RPE/sclera complex were cut into 4 pieces and digested in Liberase TL for 1 hour at 37°C with shaking (200 rpm). After digestion, single-cell suspensions were treated identically to our previously published flow cytometry procedure (22 (link)). The innate immune cell marker antibodies can be found in Table 1. Samples were run on a FACSymphony A5-Laser Analyzer (Becton Dickinson), and data were analyzed using FlowJo version 10 (FlowJo).

Droho S., Rajesh A., Cuda C.M., Perlman H, & Lavine J.A. (2023). CD11c+ macrophages are proangiogenic and necessary for experimental choroidal neovascularization. JCI Insight, 8(7), e168142.

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Immunophenotyping of Cell Surface Markers

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Approximately 1 × 10⁶ cells were incubated for 15 min with allophycocyanin (APC)-conjugated mouse anti-CD44 (BD Biosciences, 560890) and phycoerythrin ( PE)-conjugated mouse anti-CD24 (BD Biosciences, 560991) according to instructions provided by the manufacturer. Cells were analyzed using a flow cytometer instrument (BD FACSymphony A5-Laser Analyzer) equipped with FACSDiva software.

Coelho D.R., Palma F.R., Paviani V., He C., Danes J.M., Huang Y., Calado J.C., Hart P.C., Furdui C.M., Poole L.B., Schipma M.J, & Bonini M.G. (2022). Nuclear-localized, iron-bound superoxide dismutase-2 antagonizes epithelial lineage programs to promote stemness of breast cancer cells via a histone demethylase activity. Proceedings of the National Academy of Sciences of the United States of America, 119(29), e2110348119.

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Murine Lung Cell Characterization

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Mouse lung was digested and single-cell suspensions were prepared as previously described (46 (link)). Cell suspensions underwent red blood cell lysis using Pharm Lyse buffer (BD Biosciences). Live/dead staining was performed in protein-free solution (HBSS) using fixable viability dye eFluor 506 (eBioscience), followed by incubation with FcR-blocking reagent (Miltenyi Biotec). Antibodies utilized for murine cell staining included rat anti–mouse CD45–FITC (BioLegend, 30-F11), rat anti–mouse Ly6C–eFluor450 (eBiosciences, HK1.4), rat anti–mouse I-A/I-E–PerCPCy5.5 (BioLegend, M5/114.15.2), rat anti–mouse CD45–APC (BioLegend, 30-F11), rat anti–mouse Ly6G–Alexa Fluor 700 (BioLegend, 1A8), rat anti–mouse NK1.1–Alexa Fluor 700 (BD, PK136), rat anti–mouse CD11b–APCCy7 (BioLegend, M1/70), rat anti–mouse CD64–PE (BioLegend, X54-5/7.1), rat anti–mouse SiglecF–PECF594 (BD, E50-2440), and rat anti–mouse CD11c–PECy7 (BD, HL3). For neutrophil quantification, 123Count eBeads (Invitrogen) were added. Flow analysis of fixed samples was done on a BD FACSymphony A5-Laser Analyzer at the Northwestern University Robert H. Lurie Comprehensive Cancer Center Flow Cytometry Core facility. Acquired data were analyzed with FlowJo v10.8 (FlowJo).

Yang W., Cerier E.J., Núñez-Santana F.L., Wu Q., Yan Y., Kurihara C., Liu X., Yeldandi A., Khurram N., Avella-Patino D., Sun H., Budinger G.R., Kreisel D., Mohanakumar T., Lecuona E, & Bharat A. (2022). IL-1β–dependent extravasation of preexisting lung-restricted autoantibodies during lung transplantation activates complement and mediates primary graft dysfunction. The Journal of Clinical Investigation, 132(20), e157975.

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Comprehensive Immune Cell Profiling

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Single cell suspensions were incubated with blocking CD16/32 antibody (BioLegend #101301) and then Fixable Viability Dye (ThermoFisher Scientific). All staining used 0.5-5x10⁶ cells stained in 100ul 3% FBS in PBS with antibodies at manufacturer’s recommendation unless otherwise specified. Antibodies included CD4(GK1.5), CD8(53–6.7), CD25(A18246A), CD44(IM7), CD62L(MEL-14), CD45(QA17A26), CD45.1(A20), CD45.2(104), CD45RB (C363-16A), IL7Ra(A7R34), CCR7(4B12), TIGIT(1G9), ICOS (C398.4A), and GITR(DTA-1). Surface-directed antibodies were stained for 20 min at 4°C then washed before analysis or intracellular staining. For detection of intracellular proteins, the Foxp3/Transcription Factor Staining Buffer Set (Thermofisher Scientific #00-5523-00) was used. Intracellular Antibodies included FOXP3 (FJK-16s), IFNγ (XMG1.2), IL17A(TC11-18H10.1), GM-CSF(MP1-22E9), IRF8 (V3GYWCH), IRF4(IRF4.3E4), HELIOS (22F6), TCF-1(S33-966). For samples requiring intracellular cytokine staining cells were first incubated for 4 h with 25 ng/mL PMA, 0.5uM ionomycin, and 10 μg/mL monensin. All analysis was performed on a BD LSRFortessa X-20 Analyzer and a BD FACSymphony A5-Laser Analyzer. All cell sorting was performed on BD SORP FACS Aria II or Miltenyi Tyto. All analysis was performed on FlowJo v10 (BD).

Chaudhuri S.M., Weinberg S.E., Wang D., Yalom L.K., Montauti E., Iyer R., Tang A.Y., Torres Acosta M.A., Shen J., Mani N.L., Wang S., Liu K., Lu W., Bui T.M., Manzanares L.D., Dehghani Z., Wai C.M., Gao B., Wei J., Yue F., Cui W., Singer B.D., Sumagin R., Zhang Y, & Fang D. (2024). Mediator complex subunit 1 architects a tumorigenic Treg cell program independent of inflammation. Cell Reports Medicine, 5(3), 101441.

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Single-cell Analysis of Transplanted Lungs

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Lungs harvested 24 hours following orthotopic lung transplantation were processed to a single-cell suspension and were stained according to a previously described method (69 (link)). Antibodies used for lung staining can be found in Supplemental Table 1. Samples were run on a BD FACSymphony A5-Laser Analyzer at the Northwestern University Robert H. Lurie Comprehensive Cancer Center Flow Cytometry Core Facility and then subsequently analyzed with FlowJo v10.6.2 (FlowJo).

Querrey M., Chiu S., Lecuona E., Wu Q., Sun H., Anderson M., Kelly M., Ravi S., Misharin A.V., Kreisel D., Bharat A, & Budinger G.R. (2022). CD11b suppresses TLR activation of nonclassical monocytes to reduce primary graft dysfunction after lung transplantation. The Journal of Clinical Investigation, 132(14), e157262.

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