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Lsr 2 special order system

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The BD LSR II Special Order System is a flow cytometry instrument designed for advanced research applications. It features a customizable configuration to meet specific experimental requirements. The system enables users to analyze and sort cells or particles based on their physical and fluorescent properties.

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Lab products found in correlation

Propidium iodide, by Thermo Fisher Scientific (1 mentions) Rnase a, by Qiagen (1 mentions) Collagenase, by Thermo Fisher Scientific (1 mentions) 40 μm strainer, by BD (1 mentions) Facsdiva 6, by BD (1 mentions) Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions) Alexa fluor 700, by BD (1 mentions) Cyan adp, by Beckman Coulter (1 mentions)

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LSR II system BD LSR II

5 protocols using lsr 2 special order system

1

Flow Cytometry Immunophenotyping of Monocytes

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All monoclonal antibodies used for flow cytometric analysis were obtained from Becton Dickinson Immunocytometry Systems, eBioscience (San Diego, CA), and BioLegend (San Diego, CA). Each blood sample was labeled with the following 3-color antibody combinations to determine the co-expression of corresponding antigens on monocytes: APC-conjugated mouse anti-human CD45 and PE-conjugated mouse anti-human CD14; and SVF derived leukocytes were stained with mAbs (APC-Cy7-conjugated mouse anti-human CD45, PE-Cy7-conjugated mouse anti-human CD3, PE-conjugated mouse anti-human CD14, APC-conjugated mouse anti-human IL-1β) on ice for 25 min, followed by washing. Prepared blood and SVF samples were both analyzed using BD LSR II Special Order System (BD Biosciences, San Jose, CA, USA). After adjustment of the fluorochrome compensation electronically, 200,000 events per sample were acquired and analyzed. FMO controls were used to set up the gating of specific cell populations.
Jeschke M.G., Patsouris D., Stanojcic M., Abdullahi A., Rehou S., Pinto R., Chen P., Burnett M, & Amini-Nik S. (2015). Pathophysiologic Response to Burns in the Elderly. EBioMedicine, 2(10), 1536-1548.
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2

Apoptosis Induction by LEE011 and Ruxolitinib

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To measure the effect of LEE011 and ruxolitinib combinatorial treatment on apoptosis, NKYS, KHYG and NK-S1 were separately treated at 2 × 105 cells/ml in 10 cm dishes at appropriate drug concentrations, and harvested 48 h later. Cells were washed and fixed in ice-cold 70% ethanol for 2 h before further incubation at -20°C for up to 24 h. Immediately before flow cytometric analysis, the cells were treated with RNase A (Qiagen) and stained with propidium iodide (Invitrogen, USA) for 30 min. Collection and analysis of data was done using BD LSR II Special Order System (BD Biosciences, USA).
Hee Y.T., Yan J., Nizetic D, & Chng W.J. (2018). LEE011 and ruxolitinib: a synergistic drug combination for natural killer/T-cell lymphoma (NKTCL). Oncotarget, 9(61), 31832-31841.
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3

Quantification of Tissue-Resident Macrophages

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Skin was minced and digested with collagenase (Life Technologies) for 45 minutes at 37°C while shaking. The digested cell suspension was filtered and centrifuged at 1800 rpm for 5 minutes. Pelleted cells were resuspended in FACS buffer (HBSS containing 0.5% BSA) and passed through a 40μM strainer (BD Bioscience) to remove large cellular debris. Cells were stained with monoclonal antibodies on ice for 30 minutes. Samples were than washed and analyzed using BD LSR II Special Order System (BD Biosciences, San Jose, CA, USA). Cells were gated on FSC-A and SSC-A, followed by doublet exclusion (FSC-W x FSC-H, SSC-W x SSC-H). The total percentage of monocytes/macrophages was identified using the following flurochrome-conjugated antibodies: anti-CD45 (anti-mouse PE-Cyanine7, eBioscience), anti-CD11b (anti-mouse Alexa APC-eFluor® 780, eBioscience), and anti-F4/80 (anti-mouse FITC, eBioscience). Anti-inflammatory macrophages were identified using an anti-CD206 antibody (anti-mouse PE, BioLegend) in accordance with the manufacturer’s flow cytometry protocol. The gating strategy for assessing innate immune cell distributions included leukocytes initially gated based on granularity and CD45 (side scatter x CD45), followed by size (forward scatter). Gated cells were stained for cell surface markers for monocytes and macrophages (CD11b+/F4/80+).
Vinaik R., Abdullahi A., Barayan D, & Jeschke M.G. (2019). NLRP3 Inflammasome Activity is Required for Wound Healing After Burns. Translational research : the journal of laboratory and clinical medicine, 217, 47-60.
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4

PBMC Characterization by Flow Cytometry

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For each individual cell sample, 5×106 PBMCs were quickly thawed at 37°C and diluted in 40 ml cold wash buffer (PBS supplemented with 2.5% FBS (Life Technologies Ltd. Paisley, UK) and 0.1% sodium azide). The cell suspension was centrifuged at 250×g for 5 min and after discarding the supernatant, the pellet was resuspended in 400μl wash buffer. Each sample was stained for 1 h in a light-protected environment at 2°C with titrated amounts of anti CD19 labelled with Alexa Fluor 700 (BD Biosciences). Samples were analyzed using a BD LSR II Special Order System, controlled by the BD FACSDiva 6.0 software (BD Biosciences). A preliminary forward scatter (FSC) versus side scatter (SSC) gate was used to identify lymphocytes and, depending on sample size, a total of up to 100.000 in-gate events were recorded. All datasets were migrated to FlowJo 7.6.5 (Treestar Inc. Ashland, OR, USA) for further gating and analysis.
Söllvander S., Ekholm-Pettersson F., Brundin R.M., Westman G., Kilander L., Paulie S., Lannfelt L, & Sehlin D. (2015). Increased Number of Plasma B Cells Producing Autoantibodies Against Aβ42 Protofibrils in Alzheimer’s Disease. Journal of Alzheimer's Disease, 48(1), 63-72.
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5

Erythrocyte Cell Count Analysis

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Cell numbers and fluorescence intensity was assayed using either the BD LSRII Special Order System (BD, San Jose, CA, USA) or CyAn ADP (Beckman Coulter, Fort Collins, CO, USA) as described previously,6 (link) and a minimum of 100 000 erythrocytes were analyzed.
Ch'ng J.H., Lee Y.Q., Gun S.Y., Chia W.N., Chang Z.W., Wong L.K., Batty K.T., Russell B., Nosten F., Renia L, & Tan K.S. (2014). Validation of a chloroquine-induced cell death mechanism for clinical use against malaria. Cell Death & Disease, 5(6), e1305-.
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