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Facsvantage cell sorter

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Sourced in United States, Germany

The FACSVantage cell sorter is a flow cytometry instrument designed for high-performance cell sorting. It features advanced optics, fluidics, and electronics to enable accurate and efficient cell separation based on specific cellular characteristics.

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Lab products found in correlation

Cd28pe cd28.2, by BD (3 mentions) Facscalibur, by BD (3 mentions) Cellquest software, by BD (2 mentions) Cd3 percp sp 34 2, by BD (2 mentions) Facsdiva software, by BD (2 mentions) Facscan, by BD (2 mentions) Canto analyzer, by BD (2 mentions) Tryple, by Thermo Fisher Scientific (2 mentions) Facsaria 2, by BD (1 mentions) Modfit software, by Verity Software House (1 mentions) Anti human cd19 pecy7, by BD (1 mentions) Anti human cd27 pe, by BD (1 mentions) Anti human igg apc, by BD (1 mentions) Ficoll density gradient centrifugation, by GE Healthcare (1 mentions) Cd95 apc dx2, by BD (1 mentions) T cell purification columns, by R&D Systems (1 mentions) Diva enhancement software, by BD (1 mentions) Collagenase type 2, by Merck Group (1 mentions) Collagenase type 1, by Thermo Fisher Scientific (1 mentions) Rpmi 1640 medium, by Thermo Fisher Scientific (1 mentions)

38 protocols using facsvantage cell sorter

1

Plant Ploidy Level Analysis by Flow Cytometry

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Each sample of frozen cell pellet (500 mg) was treated with the High-Resolution Kit for plant ploidy level analysis [Kit Cystain UV precise type P, containing solution A (Nuclei extraction buffer) and solution B (Staining buffer containing DAPI -4,6,diamino-2-phenylindol), Partec GmbH, Munster, Germany] to determine the DNA content. To release cell nuclei, we rinsed the cells in solution A and carefully chopped with a sharp razor blade. Then, cells were filtered and rinsed in solution B17 (link). On average, 10000 particles in three replicates were counted by flow cytometry (Cell Sorter FACS Vantage, Becton–Dickinson, San Diego, California, USA), using ion laser tuned at 360 nm and detection of emission using a blue fluorescence emission filter (bandpass filter of 424/44 nm). FACS analysis results were analyzed using BD CellQuest™ software to determine the ratios of cell cycle phases per the DNA content of individual cells (2n for phase G1, 2 < n < 4 for S, 4n for G2/M phases).
Kamal K.Y., Herranz R., van Loon J.J, & Medina F.J. (2018). Simulated microgravity, Mars gravity, and 2g hypergravity affect cell cycle regulation, ribosome biogenesis, and epigenetics in Arabidopsis cell cultures. Scientific Reports, 8, 6424.
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2

DNA Content Analysis of Plant Cells

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Frozen cell pellet (500 mg) was treated with the High Resolution Kit for plant ploidy level analysis (Kit Cystain UV precise P), containing solution A (Nuclei extraction buffer) and solution B (Staining buffer containing DAPI: 4, 6, diamino-2-phenyl-indol) (Partec GmbH, Munster, Germany) to determine the DNA content. For nuclei release, the cells were rinsed in solution A, carefully chopped with a sharp razor blade, and then cells were filtered and rinsed in solution B (Menges & Murray, 2006) . On average, 10000 particles in three replicates were counted by flow cytometry (Cell sorter FACS Vantage, Becton-Dickinson, San Diego, California), using ion laser tuned at 360 nm and detection of emission using a blue fluorescence emission filter (bandpass filter of 424/44 nm Ban Pass). FACS analysis results were analyzed using BD CellQuest™ software for Windows XP to determine the ratios of cell cycle phases according to the DNA content of individual cells (2n for phase G1, 2
Kamal K.Y., Herranz R., van Loon J.J.W.A, & Medina F.J. (2019). Cell cycle acceleration and changes in essential nuclear functions induced by simulated microgravity in a synchronized Arabidopsis cell culture. Plant, cell & environment, 42(2).
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3

Cell Cycle Analysis by Flow Cytometry

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The cells were harvested and washed once with phosphate-buffered saline (PBS), fixed in pre-cold 70% ethanol and incubated with 50 µg/ml PI (Sigma-Aldrich), 1 mg/ml RNase solution, for 60 min at 4°C in the dark. The stained nuclei were analyzed with a FACS Vantage cell sorter, and the data were analyzed using a Mod-Fit 2.0 cell cycle analysis program (both from BD Biosciences). Flow cytometric analysis was performed using a BD FACSAria II flow cytometer (BD Biosciences), and the data were analyzed using ModFit software (Verity Software House, Topsham, ME, USA).
Fratangelo F., Camerlingo R., Carriero M.V., Pirozzi G., Palmieri G., Gentilcore G., Ragone C., Minopoli M., Ascierto P.A, & Motti M.L. (2018). Effect of ABT-888 on the apoptosis, motility and invasiveness of BRAFi-resistant melanoma cells. International Journal of Oncology, 53(3), 1149-1159.
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4

Multiparametric Flow Cytometry of Stem Cells

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Cells were washed with PBS and resuspended in PBS with 2.5% FBS. Cells were simultaneously stained with FITC-conjugated CD45, c-kit, and TER119 mAbs, APC-conjugated CD49f mAb, and PercpCy5.5-conjugated CD29 mAb at 4 °C for 30 min. In addition, they were incubated with PE-conjugated Dlk1 (all from eBioscience, San Diego, CA, USA). The labeled cells were analyzed using a FACSVantage cell sorter (BD, San Jose, CA, USA). Gating was implemented based on negative-control staining profiles. In some experiments, the PE-conjugated Dlk1-labeled cells were sorted into positive and negative subpopulations for further studies.
Huang J., Zhao X., Wang J., Cheng Y., Wu Q., Wang B., Zhao F., Meng L., Zhang Y., Jin M, & Xu H. (2019). Distinct roles of Dlk1 isoforms in bi-potential differentiation of hepatic stem cells. Stem Cell Research & Therapy, 10, 31.
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5

Intestinal plasmablast/plasma cell isolation

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Intestinal plasmablasts/plasma cells were isolated as previously described (Benckert et al., 2011 (link)). In brief, lamina mucosa and propria were dissected from lamina muscularis mucosae by blunt preparation, and 2–3-mm tissue pieces were digested using 0.1% DNase and 0.1% collagenase followed by discontinuous Ficoll density gradient centrifugation (GE Healthcare). Purified lamina propria lymphocytes were stained on ice with fluorochrome-coupled anti-human CD38 FITC, anti-human CD27 PE, anti-human CD19 PECy7, anti-human IgG APC (all from BD Bioscience) or anti-human IgA APC (Jackson Laboratory). Single CD38+CD27+IgA+ or CD38+CD27+IgG+ plasmablasts/plasma cells were sorted into 96-well PCR plates using a FACSVantage cell sorter with Diva configuration (BD Bioscience), snap frozen on dry ice, and stored at −80°C until further processing.
Kabbert J., Benckert J., Rollenske T., Hitch T.C., Clavel T., Cerovic V., Wardemann H, & Pabst O. (2020). High microbiota reactivity of adult human intestinal IgA requires somatic mutations. The Journal of Experimental Medicine, 217(11), e20200275.
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6

Multiparametric Flow Cytometry Analysis

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PBMCs, peripheral lymph nodes, spleen and bone marrow cells were labeled with a combination of the following mAbs: CD3 PerCP (SP 34–2), CD4 PerCP (L-200), CD8 PerCP (SK1), CD8 APC (SK1), CD95 FITC (DX2), CD95 APC (DX2), and CD28PE (CD28.2) (BD Pharmingen, San Jose, CA). For chimerism analyses, we used an anti-MHC class I HLA mAb (H38, One Lambda, Inc, CA) reacting specifically with certain MHC class I antigens. The recipient and donor pairs were chosen based on their differential reactivity to this mAb. The fluorescence of the stained samples was analyzed using FACS Calibur and FACS Scan flow cytometers and Cell Quest Software (BD), or FlowJo software. For assessing the effect of CTLA4Ig on memory T cell functions, the monoclonal antibodies anti-CD16, anti-CD95, anti-CD4 and anti-CD8 were used to purify CD4 Tmems (CD16−CD4+CD95+) and CD8 Tmems(CD16−CD8+CD95+) using a FACS Vantage cell sorter (BD Immunocytometry System), then those purified populations were tested for their IFNγ production in ELISPOT assay as previously described (11 (link), 12 (link)). The purity of sorted cells was consistently > 95% as previously described.
Yamada Y., Ochiai T., Boskovic S., Nadazdin O., Oura T., Schoenfeld D., Cappetta K., Smith R.N., Colvin R.B., Madsen J.C., Sachs D.H., Benichou G., Cosimi A.B, & Kawai T. (2014). Use of CTLA4Ig for Induction of Mixed Chimerism and Renal Allograft Tolerance in Nonhuman Primates. American journal of transplantation : official journal of the American Society of Transplantation and the American Society of Transplant Surgeons, 14(12), 2704-2712.
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7

CD31 Expression Analysis in NHAECs

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The expression of cluster of differentiation (CD)31 on NHAECs was analyzed by flow cytometry using appropriated primary and fluorescein isothiocyanate-labelled secondary antibodies (25 (link)). Samples were analyzed using a FACSCalibur/LSR II flow cytometer (BD Biosciences, Franklin Lakes, NJ, USA), and cell sorting was performed with a FACSVantage cell sorter (BD Biosciences). Data were analyzed using FACSDiva™ software (version 8.0.1; BD Biosciences). Gates were set according to isotype controls.
Amara S., Alotaibi D, & Tiriveedhi V. (2016). NFAT5/STAT3 interaction mediates synergism of high salt with IL-17 towards induction of VEGF-A expression in breast cancer cells. Oncology Letters, 12(2), 933-943.
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8

Cell Cycle Analysis by Flow Cytometry

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Cells were seeded in a six-well plate (1.5×106 cells/well) and treated with AA-PMe or AA (1–50 μM) for 24 hours. Then the cells were harvested, fixed with 75% ethanol for 2 hours at 4°C, washed with PBS, and stained with PI (5 mg/L) containing RNase A (1 g/L), sodium citrate (1 g/L), and 0.5% Triton X-100 for 30 minutes in the dark. Then the cells were analyzed by a FACS Vantage cell sorter (BD Bioscience, Franklin Lakes, NJ, USA) equipped with a 488 nm argon laser. Cell distribution in different phases of the cell cycle was estimated using ModFit LT™ software.
Jing Y., Wang G., Ge Y., Xu M., Tang S, & Gong Z. (2016). AA-PMe, a novel asiatic acid derivative, induces apoptosis and suppresses proliferation, migration, and invasion of gastric cancer cells. OncoTargets and therapy, 9, 1605-1621.
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9

Isolation and Characterization of Naïve and Memory T Cells

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T cells were isolated from the spleen and lymph nodes (axillary, inguinal and brachial) of transplanted and naïve mice by negative selection using commercially available T cell purification columns according to the manufacturer’s instructions (R & D Systems, Minneapolis, MN). Purified T cells were washed in HBSS and used in ELISPOT assays. Naïve and memory T cells were separated using a FACS Vantage cell sorter (BD Immunocytometry System), based on their expression of CD44 surface marker (CD44low: naïve T cells, CD44high: memory T cells). The purity of sorted cells was consistently > 95%. CD4+ and CD8+ T cells were isolated via FACS sorting using the following antibodies: anti-CD44 FITC (clone IM7), anti-CD8 APC (clone 54–6.7), anti-CD4 APC/Cy7 (clone GK1.5), anti-CD3 PE/Cy7 (clone 17A2).
Marino J., Gonzalez-Nolasco B., Wang X., Orent W, & Benichou G. (2020). Contrasting effects of B cell depletion on CD4+ and CD8+ memory T cell responses generated after transplantation. American journal of transplantation : official journal of the American Society of Transplantation and the American Society of Transplant Surgeons, 20(9), 2551-2558.
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10

Adoptive Transfer of OT-1 CD8+ T Cells

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Splenocytes from (OT-1-Thy1.1× A/WySnJ)F1 mice were enriched for CD8+ cells by negative selection using Ab-conjugated micromagnetic beads as described [58] (link). Cells were then stained with anti-CD44-FITC, and CD44lo cells were sorted using FACSVantage cell sorter with DIVA enhancement software (BD Bioscience). A total of 1–2×107 cells were transferred into recipient B6AF1 mice infected with either FV or FV-OVA 42 days prior to the cell transfer.
Takamura S., Kajiwara E., Tsuji-Kawahara S., Masumoto T., Fujisawa M., Kato M., Chikaishi T., Kawasaki Y., Kinoshita S., Itoi M., Sakaguchi N, & Miyazawa M. (2014). Infection of Adult Thymus with Murine Retrovirus Induces Virus-Specific Central Tolerance That Prevents Functional Memory CD8+ T Cell Differentiation. PLoS Pathogens, 10(3), e1003937.
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