Sy3200 cell sorter

Manufactured by Sony

The SY3200 cell sorter is a laboratory instrument designed for the separation and isolation of cells based on their specific characteristics. It is capable of rapidly analyzing and sorting a diverse range of cell types with high precision and efficiency.

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Lab products found in correlation

Dnase 1, by Roche (2 mentions) 100 μm cell strainer, by BD (2 mentions) Trypsin edta, by Thermo Fisher Scientific (2 mentions) Macsquant analyzer vyb, by Miltenyi Biotec (2 mentions) Hanks balanced salt solution (hbss), by Thermo Fisher Scientific (2 mentions) Cytoflex, by Beckman Coulter (2 mentions) Rneasy mini kit, by Qiagen (2 mentions) Hiseq 4000 system, by Illumina (2 mentions) Liberase, by Roche (2 mentions) Lipofectamine 2000, by Thermo Fisher Scientific (2 mentions) Icyt reflection, by Sony (2 mentions) Pgfp c shlenti, by OriGene (1 mentions) Uea 1, by Vector Laboratories (1 mentions) 7 aad viability staining solution, by BioLegend (1 mentions) Hepes, by Thermo Fisher Scientific (1 mentions) Bovine serum albumin (bsa), by Merck Group (1 mentions) Cell strainer, by Corning (1 mentions) Cd16 cd32, by BD (1 mentions) Biotinylated antibodies, by BD (1 mentions) Ld column, by Miltenyi Biotec (1 mentions)

Close spelling variants of this product

ICyt Synergy SY3200 Cell Sorter ICyt SY3200 Cell Sorter SY3200 Synergy HAPS cell sorter SY3200 sorter SY3200

31 protocols using sy3200 cell sorter

Annexin A2 Silencing in KS1767 Cells

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Silencing of annexin A2 expression was performed in KS1767 cells seeded into 6-well plates. Cell transduction was obtained with four alternative lentiviral shRNA clones directed against human annexin A2 (pGFP-C-shLenti, Origene Technologies). A scrambled shRNA lentivirus was used as a negative control. GFP-positive cells were sorted with a sy3200 Cell Sorter (Sony Biotechnology) and seeded into 6-well plates. Cell extracts were obtained in 50 mM Tris, 150 mM NaCl, 1% NP-40, phosphatase/protease inhibitors (Roche). To evaluate the downmodulation of annexin A2 protein, a Western blot was performed with a mouse monoclonal anti-annexin A2 (Novus Biologicals).

Staquicini D.I., Rangel R., Guzman-Rojas L., Staquicini F.I., Dobroff A.S., Tarleton C.A., Ozbun M.A., Kolonin M.G., Gelovani J.G., Marchiò S., Sidman R.L., Hajjar K.A., Arap W, & Pasqualini R. (2017). Intracellular targeting of annexin A2 inhibits tumor cell adhesion, migration, and in vivo grafting. Scientific Reports, 7, 4243.

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Isolation and Sorting of Thymic Epithelial Cells

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The thymi were fragmented and excess of thymocytes washed away by mechanical disruption. TECs were released by enzymatic digestion. Briefly, the thymic fragments were incubated in digestion medium (0.5 U/mL Liberase TM (Roche), 0.2 mg/mL DNase I (Roche) in DMEM/F12) at 37°C with gentle mixing for 20 min. The released cells were transferred into cold flow cytometry buffer. New pre-warmed digestion medium was added to remaining thymic fragments for two more consecutive incubations, to completely dissolve the tissue. The released cell fractions were filtered through a 100 μm cell strainer (BD Biosciences), washed and counted. Cells from the two latter fractions were pooled for analysis. After incubation with FcR block (CD16/CD32, BD Biosciences), antibodies against CD45 (30-F11, BD Biosciences) and EpCAM/CD326 (G8.8, BD Biosciences), Ly51 (6C3, BD Biosciences) and the biotinylated lectin UEA-1 (Vector Laboratories) were added. The cells were washed, resuspended in flow cytometry buffer, and filtered through a 100 μm cell strainer. mTECs (CD45⁻ EpCAM⁺ UEA1⁺ Ly51⁻) and cTECs (CD45⁻ EpCAM⁺ UEA1⁻ Ly51⁺) were sorted on a SY3200 cell sorter (SONY Biotechnology Inc.).

Wilhelmson A.S., Lantero Rodriguez M., Johansson I., Svedlund Eriksson E., Stubelius A., Lindgren S., Fagman J.B., Fink P.J., Carlsten H., Ekwall O, & Tivesten Å. (2020). Androgen Receptors in Epithelial Cells Regulate Thymopoiesis and Recent Thymic Emigrants in Male Mice. Frontiers in Immunology, 11, 1342.

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Flow Cytometry Sample Preparation

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Cells were washed by PBS, lifted
by 0.25% EDTA–Trypsin (Gibco), and diluted in HBSS (Gibco)
with 0.25% of BSA before flow cytometry. Flow cytometry experiments
were performed either on MACSQuant VYB Analyzer (Miltenyi Biotec)
or CytoFLEX (Beckman Coulter). Analysis of data was done with open
source, in-house developed software, EasyFlow (https://github.com/AntebiLab/easyflow), or EasyFlowQ (https://github.com/ym3141/EasyFlowQ).
FACS was performed
with SY3200 Cell Sorter (Sony) at Caltech FLow Cytometry Facility.

Ma Y., Budde M.W., Zhu J, & Elowitz M.B. (2023). Tuning Methylation-Dependent Silencing Dynamics by Synthetic Modulation of CpG Density. ACS Synthetic Biology, 12(9), 2536-2545.

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Isolation of Chick Embryonic Enteric Neurons

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At embryonic day 10, the gastrointestinal tract, including the associated Nerve of Remak and pelvic plexuses (Supplementary file 2), was dissected from chick embryos and washed with Ringer’s solution. Pre- and post-umbilical regions were separated, broke into pieces in chilled DPBS, and loose-fit homogenized in Accumax solution (EMD Millipore). 400 μl of Accumax-tissue mixture was aliquoted into 1.7 ml Eppendorf tubes and shaken at 37°C for 12 min. After dissociation, chilled Hanks Buffered Saline Solution (HBSS) supplemented by BSA (125 mg in 50 ml, Sigma; 0.2% w/v) and 1 M HEPES (500 μl in 50 ml, pH 7.5, Thermo Fisher) was added to quench the reaction. The dissociated cells were passed through a 70 μm cell strainer (Corning) and collected by centrifuging at 500 × g for 11 min at 4°C. The cells were resuspended in HBSS-BSA, supplemented 7-AAD Viability Staining Solution (BioLegend # 420404, 500 TESTS), and sorted for YFP+, viable single cells using Sony SY3200 cell sorter at the Caltech Flow Cytometry Facility.

Jacobs-Li J., Tang W., Li C, & Bronner M.E. (2023). Single-cell profiling coupled with lineage analysis reveals vagal and sacral neural crest contributions to the developing enteric nervous system. eLife, 12, e79156.

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Sorting Transfected Neuroblastoma Cells

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GFP positive and negative UKF-NB-MV transfected cells were sorted sterilely with a SY3200 Cell Sorter (Sony Biotechnology, Inc.) and collected for further growth. We obtained two subpopulations: GFP positive cells transfected with GFP-miRNA-124 or GFP-control plasmids (see below). Single cell suspensions were prepared (2 x 10⁶ cells in 500 µl of sterile PBS). A GFP negative control (mock-transfected UKF-NB-MV) was used to detect auto-fluorescence. The cells were collected into BD Falcon 12x75 mm polystyrene tubes coated with 4% BSA. The sorting media was RPMI-1640 supplemented with 10% heat-inactivated fetal calf serum, 1% L-glutamine, and 1% pen-strep). Cells were sorted two days after transfection where >90% of the cells were GFP positive. 30,000 cells/well of each type were collected and plated in 6-well plates.

Naaman H., Rall G., Matullo C., Veksler-Lublinsky I., Shemer-Avni Y, & Gopas J. (2017). MiRNA-124 is a link between measles virus persistent infection and cell division of human neuroblastoma cells. PLoS ONE, 12(10), e0187077.

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Isolation and Purification of Thymic Epithelial Cells

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The thymi were fragmented and excess of thymocytes washed away by mechanical disruption. TECs were released by enzymatic digestion. Briefly, the thymic fragments were incubated in digestion medium: 0.5 U/mL Liberase (Cat No. 5401127001; Roche), 0.2 mg/mL DNase I (Cat No. 11284932001; Roche) in DMEM/F12 at 37°C with gentle mixing for 20 minutes. The released cells were transferred into cold flow cytometry buffer. New prewarmed digestion medium was added to remaining thymic fragments for 2 more consecutive incubations, to completely dissolve the tissue. The released cell fractions were filtered through a 100-μm cell strainer (BD Biosciences), washed, and counted. Cells from the 2 latter fractions were pooled and used for cell sorting. After incubation with FcR block (CD16/CD32; BD Biosciences; 1:100, 5 µg/mL), antibodies against CD45 (30-F11; BD Biosciences; 1:200, 0.5 µg/mL) and EpCAM (epithelial cellular adhesion molecule)/CD326 (G8.8; BD Biosciences; 1:300, 0.7 µg/mL) were added. The cells were washed, resuspended in flow cytometry buffer to 10⁷ per mL, and filtered through a 100-μm cell strainer. TECs (CD45⁻ EpCAM⁺) were sorted on a SY3200 cell sorter (SONY Biotechnology, Inc).

Wilhelmson A.S., Lantero Rodriguez M., Svedlund Eriksson E., Johansson I., Fogelstrand P., Stubelius A., Lindgren S., Fagman J.B., Hansson G.K., Carlsten H., Karlsson M.C., Ekwall O, & Tivesten Å. (2018). Testosterone Protects Against Atherosclerosis in Male Mice by Targeting Thymic Epithelial Cells—Brief Report. Arteriosclerosis, Thrombosis, and Vascular Biology, 38(7), 1519-1527.

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Purification and Proliferation of OT-I Cells

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Total spleen CD8+ OT-I cells for in vivo assays were purified from CD45.1 OT-I transgenic mice by negative selection as follows: 30 min incubation of spleen single cell suspensions with biotinylated antibodies (all BD Biosciences) for anti-mouse CD16/CD32 (clone 2.4G2), CD4 (clone GK1.5), B220 (clone RA3-6B2), CD11c (clone HL3), CD11b (clone M1/70), Gr-1 (clone RB6-8C5) and I-A/I-E (MHC-II, clone 2G9) at 4 °C shaking, washing, 20 min incubation with Streptavidin MicroBeads in autoMACS™ Running Buffer and magnetic separation using LD columns (all Miltenyi Biotec) according to manufacturer’s instructions. For ex vivo proliferation assay, naive CD8+ CD44- CD62L+ OT-I cells were isolated by flow cytometric sorting using the SY3200 Cell Sorter (Sony Biotechnology). Cells were labeled with CellTrace™ Violet Cell Proliferation Kit (Thermofisher, Molecular Probes) according to manufacturer’s instructions. 1-2 × 10⁵ total labeled OT-I cells in 100 μl PBS were injected intravenously into mice. Naive OT-I cells were cultured for 3 days with pre-treated cDC1s in R10 medium (1:1 ratio) in round-bottom 96-well plates at 37 °C in 5% CO₂ followed by flow cytometric analysis of proliferation by dilution of the CellTrace™ Violet dye.

Wculek S.K., Amores-Iniesta J., Conde-Garrosa R., Khouili S.C., Melero I, & Sancho D. (2019). Effective cancer immunotherapy by natural mouse conventional type-1 dendritic cells bearing dead tumor antigen. Journal for Immunotherapy of Cancer, 7, 100.

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Single-cell isolation and sorting

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Single cell suspensions from digested bones and lungs were first enriched for PyMT-Bo1 tumor cells using Thy1.1 MACS beads (Miltenyi Biotec) according to the manufacturer protocol. After MACS enrichment, cells were blocked with 1:200 anti-mouse CD16/32 (2.4G2) for 10 min and then stained with 1:200 anti-mouse CD45 (30-F11, PacBlue) and 1:200 anti-mouse Thy1.1 (OX-7, PE-Cy7) in FACS buffer (0.5% BSA in PBS with 2 mM EDTA). 0.1 µg/ml DAPI (Sigma) was added to the sample 5 min before FACS analysis to stain for dead cells. PyMT-Bo1 cells were sorted into 96-well plates containing 2 µl of 10 × Lysis buffer (Clontech) with 5% RNase inhibitor (Clontech), 1 cell per well. Plates were then sealed, spun and stored in − 80 °C before proceeding to scRNA-seq. A Sony SY3200 cell sorter was used for single cell sorting.

Ren Q., Khoo W.H., Corr A.P., Phan T.G., Croucher P.I, & Stewart S.A. (2022). Gene expression predicts dormant metastatic breast cancer cell phenotype. Breast Cancer Research : BCR, 24, 10.

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Transcriptional Profiling of GFP-SOX10 in MN1 Cells

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The human SOX10 open-reading frame in pDONR221 (Addgene plasmid #24749) was obtained and SOX10 was cloned into pcDNA-DEST53 using LR Clonase (ThermoFisher) to generate a GFP-SOX10 expression construct. MN1 cells [29 (link)] were plated at a density of 100,000 cells per mL and transfected with the GFP-SOX10 expression construct using Lipofectamine 2000 (ThermoFisher). After 48 h, cells were harvested, suspended in PBS, and sorted into GFP-positive and GFP-negative populations using a SY3200 Cell Sorter (Sony). RNA was isolated from each cell population using the RNeasy Mini Kit (QIAGEN). A CAGE library was generated for each sample as described [47 (link)], with modifications made for compatibility with current generation Illumina sequencing platforms. Each library was sequenced on an Illumina MiSeq (4 million reads generated per library). The sequencing data was analyzed using FastQC (http://www.bioinformatics.babraham.ac.uk/projects/fastqc), filtered using TagDust2 [68 (link)], mapped using BWA [69 (link)], clustered using Paraclu [70 (link)], and visualized using BEDtools [71 (link)] and the UCSC Genome Browser [23 (link)].

Law W.D., Fogarty E.A., Vester A, & Antonellis A. (2018). A genome-wide assessment of conserved SNP alleles reveals a panel of regulatory SNPs relevant to the peripheral nerve. BMC Genomics, 19, 311.

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Isolation of Epidermal Melanocytes and Keratinocytes

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Wound scar tissue was harvested at day 17 after wounding. Wound tissue (lack hair follicles) was carefully cut out from wound periphery with scissors. The connective tissues were carefully removed with forceps. The wound tissue was incubated in 0.25% Trypsin (Invitrogen) for 2 hours at 37° C. Wound epidermis was separated from the dermis using forceps and scalpel blades and the epidermis was chopped finely and transferred into Media A (DMEM, 10% FBS, 1x penicillin/streptomycin). The epidermal melanocyte and keratinocyte mixture was stirred at RT for 20 min to generate single cells. The obtained single cell suspension was filtered through a 70μm nylon filter and centrifuged at 200 g for 7 min and re-suspended in PBS+10% FBS. The cell suspension was incubated with PE-Cy7-anti-mouse CD117 (1:300, BD Pharmingen) for 15 minutes at room temperature. Cells were pelleted and washed twice in PBS+10% FBS then incubated with APC-CY7-anti-mouse CD45 (1:300, BD Pharmingen) for 15 minutes at room temperature. Cells were washed and sorted on a SY3200 cell sorter (Sony Biotechnology Inc.). Epidermal melanocytes were collected as PE-Cy7 positive and APC-Cy7 negative population (Kawaguchi et al., 2008 (link)). Epidermal keratinocytes were collected as PE-Cy7 negative and APC-Cy7 negative population.

Sun Q., Rabbani P., Takeo M., Lee S.H., Lim C.H., Noel E.N., Taketo M.M., Myung P., Millar S, & Ito M. (2018). Dissecting Wnt signaling for melanocyte regulation during wound healing. The Journal of investigative dermatology, 138(7), 1591-1600.

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