Cyan immunocytometry system

Manufactured by Agilent Technologies

Sourced in Denmark

The CyAn immunocytometry system is a flow cytometry instrument designed for the detection and analysis of cells and particles in a liquid sample. It is capable of simultaneously measuring multiple parameters, including cell size, granularity, and the expression of specific surface markers or intracellular molecules.

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Lab products found in correlation

Lsrfortessa, by BD (1 mentions) Attune nxt flow cytometer, by Thermo Fisher Scientific (1 mentions) Annexin 5 staining kit, by Thermo Fisher Scientific (1 mentions) Facsaria sorp sorter, by BD (1 mentions) Fixation permeabilization solution, by BD (1 mentions) Percp conjugated antibodies to mouse cd45, by BD (1 mentions) Allophycocyaninconjugated anti mouse cd11b, by Abcam (1 mentions) Secondary goat anti rabbit fitc, by Santa Cruz Biotechnology (1 mentions) Secondary goat anti mouse fitc, by Santa Cruz Biotechnology (1 mentions) Facsaria 3 sorter, by BD (1 mentions) H2 kb, by BD (1 mentions) Dapi dilactate, by Merck Group (1 mentions)

Spelling variants of this product

3-laser CyAn immunocytometry system (3 citations)

5 protocols using cyan immunocytometry system

Annexin V Flow Cytometry Protocol

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All the antibody information is provided in supplementary table 1. Annexin V were detected by Annexin V staining kit from ebioscience (catalog# 88-8103-74) according to manufacturer’s protocol. Flow cytometry analyses were performed with CyAn Immunocytometry system (DAKO Cytomation, Fort Collins, CO), Attune NxT Flow Cytometer (ThermoFisher Scientific) and BD LSRFortessa (Franklin Lakes, NJ), the resulting data were analyzed with FlowJo software (Tree Star, Ashland, OR). Cell sorting was performed with a BD FACS Aria SORP sorter at the City of Hope FACS facility. The sorted cells were used for transfer experiments.

Song Q., Wang X., Wu X., Kang T.H., Qin H., Zhao D., Jenq R.R., van den Brink M.R., Riggs A.D., Martin P.J., Chen Y.Z, & Zeng D. (2021). IL-22-dependent dysbiosis and mononuclear phagocyte depletion contribute to steroid-resistant gut graft-versus-host disease in mice. Nature Communications, 12, 805.

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Tumor-Associated Myeloid Cell Profiling

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For in vivo staining, tumor tissue was minced and digested with trypsin for 20 minutes at 37°C. Tissue homogenate was then filtered through a 40 μm filter and prepped using Fixation/Permeabilization solution according to the manufacturer's instructions (BD Pharmingen. San Diego, CA). Multiple-color FACS analyses was performed at City of Hope FACS facility using a 3-laser CyAn immunocytometry system (Dako Cytomation, Fort Collins, CO), and data was analyzed using FlowJo software (TreeStar, San Carlos, CA) as described before (19 (link)). PerCP-conjugated antibodies to mouse CD45 (Cat: 557235) was purchased from BD Pharmingen (San Diego, CA) Allophycocyaninconjugated anti-mouse CD11b (Cat: 17-0112-82) and the primary rabbit-anti-mouse RAGE (Cat: ab3611-100) were purchased from Abcam (Middlesex, NJ). Secondary goat anti-rabbit-FITC (Cat: sc-2012) and secondary goat anti-mouse-FITC (Cat: sc-2010) were purchased from Santa Cruz Biotechnology (Santa Cruz, CA). Tumor-associated MPs (CD11b^high CD45^high) and MG (CD11b^high CD45^low) were distinguished based on previously described FACS analysis (21 (link)).

Chen X., Zhang L., Zhang I.Y., Liang J., Wang H., Ouyang M., Wu S., da Fonseca A.C., Weng L., Yamamoto Y., Yamamoto H., Natarajan R, & Badie B. (2014). RAGE Expression in Tumor-associated Macrophages Promotes Angiogenesis in Glioma. Cancer research, 74(24), 7285-7297.

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Purification and Analysis of Donor CD4+ T Cells

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Monoclonal antibodies to mouse CD4, CD8, and H-2k^b, were all purchased from BD Pharmingen (San Diego, CA). Multiple-color fluorescence-activated cell sorter (FACS) analysis was performed at City of Hope FACS facility using a 3-laser CyAn immunocytometry system (Dako Cytomation, Fort Collins, CO), and data were analyzed using FLOWJO software (Tree Star, San Carlos, CA) as previously described (36 (link)). Donor CD4⁺ T cells were stained with donor marker H-2k^b anti-mouse CD4⁺ and sorted using BD FACSAria™ III sorter(BD Biosciences, San Jose, CA) at City of Hope FACS facility. The sorted cells were used for total RNA and protein isolation.

Deng R., Cassady K., Li X., Yao S., Zhang M., Racine J., Lin J., Chen L, & Zeng D. (2014). B7H1/CD80 interaction augments PD-1-dependent T cell apoptosis and ameliorates graft versus host disease. Journal of immunology (Baltimore, Md. : 1950), 194(2), 560-574.

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Multiparameter Flow Cytometry Analysis

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PE CD4 (GK1.5), FITC CD5.1 (H11-86.1), FITC B220 (RA3-6B2), FITC H-2K^b (AF6-88.5) and Pacific Blue CD45R were purchased from BD Pharmingen (San Diego, CA). APC-CD8a (53-6.7), APC-eFluor 780 TCRβ (H57-597), PE-Cy7 TCRβ (H57-597) were purchased from eBioscience (San Diego, CA). Alexa Fluor® 488 Goat Anti-Mouse IgG (H+L), Streptavidin Alexa Fluor® 555 Alexa Fluor® 488 Donkey Anti-Rat IgG (H+L) were purchased from Life Technologies (Grand Island, NY). DAPI dilactate was purchased from Sigma (St. Louis, MO). Biotinylated Ulex europaeus agglutinin (UEA) I was purchased from Vector Laboratories (Burlingame, CA). Aqua fluorescent reactive dye for viability analysis was purchased from Invitrogen (Carlsbad, CA). APC CD20 (18B12) was the kind gift of David Serreze (Jackson Laboratory, Bar Harbor, ME) [54 (link)]. Flow cytometry experiments were performed and run on a CyAn Immunocytometry system (DAKO Cytomation, Fort Collins, CO) and the resulting data were analyzed with FlowJo software (Tree Star, Ashland, OR) [21 , 22 , 64 (link)]. Purified anti-CD20 mAb (isotype IgG2a, 5D2) was the kind gift of Genentech (San Francisco, CA).

Johnston H.F., Xu Y., Racine J.J., Cassady K., Ni X., Wu T., Chan A., Forman S, & Zeng D. (2014). Administration Of Anti-CD20 mAb Is Highly Effective In Preventing But Ineffective In Treating Chronic GVHD While Preserving Strong GVL Effects. Biology of blood and marrow transplantation : journal of the American Society for Blood and Marrow Transplantation, 20(8), 1089-1103.

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Multiparameter Flow Cytometry Analysis

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The following anti-mouse mAbs were purchased from BD Biosciences Pharmingen (San Jose, CA), eBioscience (San Diego, CA), BioLegend (San Diego,CA) or R&D Systems (Minneapolis, MN): TCRβ (H57-597), CD4 (RM4-5), CD8α (53-6.7), CD11c (HL3), CD103 (M290), α4β7 (DATK32), CCR9 (CW-1.2), CD45.2 (104), CCR7(4B12), Fas/CD95 (554258), FasL/CD178 (MFL3), CD49b (DX5), CD90.2/Thy1.2 (30-H12), CD19 (eBio1D3), Annexin V (17-8007-72). Fluorescence-activated cell sorting (FACS) was performed with a 4-laser MoFlo Immunocytometry System (Dako, Glostrup, Denmark) or CyAn immunocytometry system (Dako Cytomation, Fort Collins, CO), and data were analyzed with FlowJo software (TreeStar, Ashland, OR), as described previously [40 (link)].

He W., Racine J.J., Johnston H.F., Li X., Li N., Cassady K., Liu C., Deng R., Martin P., Forman S, & Zeng D. (2014). Depletion of host CCR7+ Dendritic Cells Prevented Donor T cell Tissue Tropism in Anti-CD3-Conditioned Recipients. Biology of blood and marrow transplantation : journal of the American Society for Blood and Marrow Transplantation, 20(7), 920-928.

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