Facsaria fusion instrument

Manufactured by BD

Sourced in United States, Canada

The BD FACSAria Fusion instrument is a high-performance cell sorter designed for advanced research applications. It features a modular, flexible design and offers multiple laser and detector options to accommodate a wide range of fluorescent markers. The instrument enables accurate cell sorting and high-resolution data acquisition for diverse research needs.

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Lab products found in correlation

Lsrfortessa, by BD (3 mentions) Facsdiva software, by BD (3 mentions) Axioimager microscope, by Zeiss (2 mentions) Giemsa staining, by Merck Group (2 mentions) Easysep human memory cd4 t cell enrichment kit, by STEMCELL (2 mentions) Lipofectamine 2000 reagent, by Thermo Fisher Scientific (2 mentions) Penicillin, by Thermo Fisher Scientific (2 mentions) Streptomycin, by Thermo Fisher Scientific (2 mentions) 4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (2 mentions) Fetal bovine serum (fbs), by Thermo Fisher Scientific (2 mentions) Cd3 percp clone sk7, by BioLegend (2 mentions) Cd4 apc clone okt4, by BioLegend (2 mentions) Cd8 bv570 clone rpa t8, by BioLegend (2 mentions) Cd14 fitc clone 63d3, by BioLegend (2 mentions) Cd19 bv510 clone sj25c1, by BD (2 mentions) Ghost dye a710 viability stain, by Cytek Biosciences (2 mentions) Lobind plate, by Eppendorf (2 mentions) Brilliant stain buffer, by BD (2 mentions) 4 6 diamidino 2 phenylindole (dapi), by Merck Group (2 mentions) Hoechst 33342, by Thermo Fisher Scientific (2 mentions)

Spelling variants of this product

FACSAria (4019 citations) FACSAria Fusion (833 citations) FACSAria instrument (57 citations) BD FACSAria Fusion flow cytometry instrument (2 citations) FACSAria Fusion Special Order Instrument (2 citations)

36 protocols using facsaria fusion instrument

Multiparametric Analysis of Immune Cells

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Flow cytometry and cell sorting were completed on a FACSAria Fusion instrument (BD) and analyzed using FlowJo analysis software (Tree Star). Surface staining was performed at 4°C in the presence of Fc block (2.4G2) in magnetic-activated cell sorting (MACS) buffer (PBS, 0.5% BSA, 2 mM EDTA). Intracellular IRF8 staining was performed using the Foxp3 staining kit (eBioscience 00-5523-00).
The following antibodies were used: CD19 (1D3), CD135 (A2F10.1), MHCII (M5/114.15.2), CD117 (2B8), B220 (RA3-6B2), CD3 (145-2C11), and CD4 (RM4-5) from BD Biosciences; CD3 (145-2C11), CD4 (GK1.5), and MHCII (M5/114.15.2) from Tonbo Biosciences; TER-119 (TER-119), Ly-6G (1A8), B220 (RA3-6B2), CD24 (M1/69), CD115 (AFS98), XCR1 (ZET), CD19 (6D5), CD8α (53-6.7), CD4 (RM4-5), CD11c (N418), and CD3 (17A2) from Biolegend; CD105 (MJ7/18), Siglec-H (eBio440c), CD3 (17A2), CD8α (53-6.7), CD11c (N418), and IRF8 (V3GYWCH) from eBiosciences; and SA-Qdot 605, CD11c (N418), CD317 (eBio927), CD172a (P84), and TCRβ (H57-597) from Invitrogen.

Liu T.T., Ou F., Belk J.A., Bagadia P., Anderson DA I.I.I., Durai V., Yao W., Satpathy A.T., Murphy T.L, & Murphy K.M. (2023). Cis interactions in the Irf8 locus regulate stage-dependent enhancer activation. Genes & Development, 37(7-8), 291-302.

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Immunostaining and FACS Analysis of Fetal Liver Cells

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Fetal liver cells were immunostained with two separate panels (see Supplementary Table 15 for antibody details). Cells were stained for 30 minutes on ice followed by DAPI staining. FACS was performed on a BD FACSAria™ Fusion instrument, and data analysed using FlowJo (v10.4.1). Cells were isolated into chilled FACS tubes coated with FBS and prefilled with 500μL sterile PBS for cytospin (500 – 2000 cells), or into 1.5mL microfuge tubes containing 20μL lysis buffer (100 cells). Giemsa staining (Sigma-Aldrich) was used to determine the morphology of sorted cells on cytospins. Slides were viewed using a Zeiss AxioImager microscope, images taken of 4 fields from n = 3 samples using the 100x objective, and viewed using Zen (v2.3). Raw images supplied in Supplementary Images.

Popescu D.M., Botting R.A., Stephenson E., Green K., Webb S., Jardine L., Calderbank E.F., Polanski K., Goh I., Efremova M., Acres M., Maunder D., Vegh P., Gitton Y., Park J.E., Vento-Tormo R., Miao Z., Dixon D., Rowell R., McDonald D., Fletcher J., Poyner E., Reynolds G., Mather M., Moldovan C., Mamanova L., Greig F., Young M., Meyer K.B., Lisgo S., Bacardit J., Fuller A., Millar B., Innes B., Lindsay S., Stubbington M.J., Kowalczyk M.S., Li B., Ashenbrg O., Tabaka M., Dionne D., Tickle T.L., Slyper M., Rozenblatt-Rosen O., Filby A., Carey P., Villani A.C., Roy A., Regev A., Chedotal A., Roberts I., Göttgens B., Behjati S., Laurenti E., Teichmann S.A, & Haniffa M. (2019). Decoding human fetal liver haematopoiesis. Nature, 574(7778), 365-371.

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Immunostaining and FACS Analysis of Fetal Liver Cells

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Isolation and Transfer of Memory CD8+ T Cells

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Single-cell suspensions were obtained by mincing spleens with a scalpel and then by mashing them through a 100 μm nylon cell strainer (BD Falcon). Red blood cells were lysed with hypotonic ammonium-chloride-potassium (ACK) lysis buffer. The mouse CD8⁺ T cell-enrichment kit (Miltenyi Biotech) was used for CD8 T cell-isolation. Memory OT-I T cells were generated by transferring low numbers of naive OT-I T cells into CD45.1 congenic host mice and by infecting the hosts with 1000 CFU of Lm-N4. Memory cells were re-isolated via staining and positive selection of live cells in 2% FCS RPMI medium (Sigma Aldrich). First, cells were stained with fluorescein isothiocyanate (FITC)-KLRG1, or phycoerythrin (PE)-CD127, and biotin-conjugated CD45.2 antibodies, and then enriched via MACS separation using anti-biotin MicroBeads (Miltenyi Biotech), in accordance with the manufacturer’s instructions. Afterward, cells were sorted for KLRG1⁺ or CD127⁺ populations using a FACS Aria Fusion instrument (BD) and then transferred into new CD45.1/1-congenic C57BL/6J host mice.

Flosbach M., Oberle S.G., Scherer S., Zecha J., von Hoesslin M., Wiede F., Chennupati V., Cullen J.G., List M., Pauling J.K., Baumbach J., Kuster B., Tiganis T, & Zehn D. (2020). PTPN2 Deficiency Enhances Programmed T Cell Expansion and Survival Capacity of Activated T Cells. Cell Reports, 32(4), 107957.

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Inducible Overexpression of TSHZ2 in Breast Cancer Cells

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siRNA-TSHZ2 (ON-TARGETplus; Dharmacon) or siRNA-CTRL (ON-TARGETplus Non-targeting; Dharmacon) was transfected into cells using the Dharmafect-1 reagent. pCMV-MYC-TSHZ2 and pDsRed-TSHZ2 were gifts from K. Kasai (Aichi Medical University School of Medicine). pEGFP-PRC1 was a gift from K. Hui (National Cancer Centre, Singapore). pEGFP-KIF4A was a gift from K. Samejima (University of Edinburgh). pEGFP-ACTN1 (#11908) and pSpCas9(BB)-2A-Puro (PX459) V2.0 (#62988) were from Addgene. Transient transfections were performed using Lipofectamine 2000 Reagent (Thermo Fisher Scientific), or jetPEI (Polyplus-transfection). The lentiviral vector pLV[Tet]-EGFP-TRE3G>hTSHZ2 (Vector-Builder, ID: VB180711-1040ara) was used for inducible overexpression of TSHZ2 (iTSHZ2), and the pHIV-Luc-ZsGreen (Addgene, #39196) was used to generate GFP-labeled cells. Viral particles encoding iTSHZ2, ZsGreen plasmid, or shRNAs targeting TSHZ2 (Sigma-Aldrich) were produced in HEK293T cells using the vectors psPAX2 (Addgene, #12260) and pMD2.G (Addgene #12259). HCC70, MDA-MB-231, and 4T1 cells were infected and selected under puromycin (2 μg/ml) or sorted using FACSAria Fusion instrument (BD Bioscience). Inducible overexpression experiments used doxycycline (2 μg/ml). For stable overexpression of TSHZ2, we used pDsRed-TSHZ2 (Takara, #632466), along with Amaxa Cell Line Nucleofector Kit V (Lonza).

Uribe M.L., Dahlhoff M., Batra R.N., Nataraj N.B., Haga Y., Drago-Garcia D., Marrocco I., Sekar A., Ghosh S., Vaknin I., Lebon S., Kramarski L., Tsutsumi Y., Choi I., Rueda O.M., Caldas C, & Yarden Y. (2021). TSHZ2 is an EGF-regulated tumor suppressor that binds to the cytokinesis regulator PRC1 and inhibits metastasis. Science signaling, 14(688), eabe6156.

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Expansion and Isolation of Ara h 1-Specific CD4+ T Cells

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Cryopreserved PBMCs from 1 of the 27 individuals (patient 107) were thawed and cultured in RPMI 1640 supplemental with 2 mM Glutamax (both from Gibco), 10% human serum (MilliporeSigma), and 100 U/ml penicillin and 100 μg/ml streptomycin (Thermo Fisher) (complete RPMI), with 50 μg/ml natural Ara h 1 (Indoor Biotechnologies), at a density of 6 × 10⁶ cells in 1 ml medium per well in 24-well plates. Complete RPMI + 10 U/mL IL-2 (R&D Systems) was added after 5 days, and cells were cultured for a total of 14 days to expand Ara h 1–specific CD4⁺ T cells. After harvesting, memory CD4⁺ T cells were isolated with the EasySep human memory CD4⁺ T cell enrichment kit (Stemcell Technologies) and labeled with APC-conjugated Ara h 1 (DRB1*03:01, amino acid 415–425) tetramer (32 (link)) (made in-house), at a concentration of 10 nM for 1 hour at room temperature. After washing off excess tetramer, the cells were labeled with BUV395-conjugated anti-CD3 (clone UCHT1; BD Biosciences), APC-Cy7–conjugated anti-CD4, FITC-conjugated anti-CD45RA, and Live/Dead Fixable Blue stain (L23105; Thermo Fisher) for 30 minutes at 4°C. Live CD3⁺CD4⁺CD45RA^– tetramer⁺ and tetramer^– T cells were sorted with a FACSAria Fusion instrument (BD Biosciences), and genomic DNA was isolated using the AllPrep DNA/RNA Micro Kit.

Smith N.P., Ruiter B., Virkud Y.V., Tu A.A., Monian B., Moon J.J., Love J.C, & Shreffler W.G. (2021). Identification of antigen-specific TCR sequences based on biological and statistical enrichment in unselected individuals. JCI Insight, 6(13), e140028.

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Flow Cytometric Separation of Clones

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We used the scRNA-seq data to identify cell surface markers that are differentially expressed between co-existing clones, in order to physically separate them via flow cytometry. For fluorescence-activated cell sorting (FACS), cells were incubated for 30 min on ice with antibodies at dilutions determined by titration experiments. Antibodies used in this study include: anti-human CD13 (ANPEP) PeCy7 (clone WM15; BioLegend) 1:10, anti-human CD184 (CXCR4) BV421 (clone 12G, BioLegend) 1:10, anti-human TM4SF4 APC (R&D) 1:10, anti-human ITM2C (clone 2E8G11, Proteintech) 1:10. For the detection of the unconjugated anti-human ITM2C antibody cells were subsequently washed and stained for 30 min on ice with anti-mouse IgG2A FITC (Biolegend) 1:100. Corresponding isotype immunoglobulin served as controls. Flow cytometric sorting was performed using a FACSAriaFusion instrument (BD Biosciences, San Jose, CA, USA) and analyses were performed using the FlowJo software (Tree Star, Ashland, OR, USA).

Andor N., Lau B.T., Catalanotti C., Sathe A., Kubit M., Chen J., Blaj C., Cherry A., Bangs C.D., Grimes S.M., Suarez C.J, & Ji H.P. (2020). Joint single cell DNA-seq and RNA-seq of gastric cancer cell lines reveals rules of in vitro evolution. NAR Genomics and Bioinformatics, 2(2), lqaa016.

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Mitochondrial Mass Analysis of Kidney Organoids

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Developing kidney organoids on day 14 of differentiation were cultured in EGM or REGM media for 4 additional days. In order to assess mitochondrial mass, organoids were stained with MitoTracker Green FM (100 nM), a mitochondrial specific fluorescent dye at 37°C for 30 min. After incubation, kidney organoids were washed twice with PBS and disaggregated into single cell suspension using AccumaxTM for 10 minutes followed by Trypsin-EDTA 0.25% (ThermoFisher) incubation for at least 10 minutes at 37°C. Once cells dissociated, FACS buffer (PBS supplemented with 5% of FBS) was added to cease the trypsin activity and samples were centrifuged for 5 minutes at 1800 rpm. After removing the supernatant, the pellet was resuspended in 300 μl of FACS buffer and the suspension was filtered into FACS tubes. Nuclei were stained with DAPI (ThermoFisher). Cells were counted using FACS Aria Fusion Instrument (BD Biosciences). FlowJo software version 10 was used for data analysis.

Dhillon P., Park J., del Pozo C.H., Li L., Doke T., Huang S., Zhao J., Kang H.M., Shrestra R., Balzer M.S., Chatterjee S., Prado P., Han S.Y., Liu H., Sheng X., Dierickx P., Batmanov K., Romero J.P., Prósper F., Li M., Pei L., Kim J., Montserrat N, & Susztak K. (2020). The nuclear receptor ESRRA protects from kidney disease by coupling metabolism and differentiation. Cell metabolism, 33(2), 379-394.e8.

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Staining and Analysis of Kidney Organoids

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Kidney organoids were stained with fluorescein-conjugated LTL (FL-1321, Vector Laboratories) as described elsewhere (Garreta et al., 2019 (link)). Kidney organoids were then dissociated to single cells using Accumax (07921, Stem Cell Technologies) for 15 min followed by 0.25% (wt/vol) trypsin (25300–054, Life Technologies) for 15 min at 37 °C. SA3800 software version 2.0.4 (SONY) was used to acquire flow cytometry samples in the Sony SA3800 spectral cell analyzer (SONY). FACSDiva software version 8.0.1 (BD Biosciences) was used in the FACS Aria Fusion instrument (BD Biosciences) for cell sorting experiments. FlowJo software version 10 was used to analyze the data.

Garreta E., Prado P., Stanifer M.L., Monteil V., Marco A., Ullate-Agote A., Moya-Rull D., Vilas-Zornoza A., Tarantino C., Romero J.P., Jonsson G., Oria R., Leopoldi A., Hagelkruys A., Gallo M., González F., Domingo-Pedrol P., Gavaldà A., del Pozo C.H., Hasan Ali O., Ventura-Aguiar P., Campistol J.M., Prosper F., Mirazimi A., Boulant S., Penninger J.M, & Montserrat N. (2022). A diabetic milieu increases ACE2 expression and cellular susceptibility to SARS-CoV-2 infections in human kidney organoids and patient cells. Cell Metabolism, 34(6), 857-873.e9.

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Isolation and Purification of Cardiac Nuclei

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Cardiac tissues were thawed on ice and cut into small pieces. Minced tissue was pre-digested with a 5-ml enzyme solution of collagenase (2500 U, Thermo Fisher Scientific) in HBSS + /+ (Gibco) for 10 min at 37 °C in a water bath. After centrifugation at 500 g and 4 °C for 5 min, the supernatant was discarded, and nuclei were isolated after cell disruption with a glass dounce homogenizer (five strokes with a loose pestle and ten strokes with a tight pestle). After filtering (20-µm strainer, pluriSelect), the suspension was centrifuged at 1000 g and 4 °C for 6 min and resuspended in 500 µl staining buffer containing 1% BSA (Sigma-Aldrich), 5 nM MgCl2 (Sigma-Aldrich), 1 mM EDTA (Gibco), 1 mM EGTA (Gibco), 0.2 U µl−1 RNasin Plus Inhibitor (Promega) and 0.1 µg ml−1 Hoechst (Life Technologies) in Dulbecco´s phosphate buffered saline (DPBS). Hoechst-positive nuclei were separated from cell debris by using the FACSAria Fusion instrument (BD Biosciences) and sorted into a staining buffer without Hoechst at 4 °C.

Shumliakivska M., Luxán G., Hemmerling I., Scheller M., Li X., Müller-Tidow C., Schuhmacher B., Sun Z., Dendorfer A., Debes A., Glaser S.F., Muhly-Reinholz M., Kirschbaum K., Hoffmann J., Nagel E., Puntmann V.O., Cremer S., Leuschner F., Abplanalp W.T., John D., Zeiher A.M, & Dimmeler S. (2024). DNMT3A clonal hematopoiesis-driver mutations induce cardiac fibrosis by paracrine activation of fibroblasts. Nature Communications, 15, 606.

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