Facscanto analyzer

Manufactured by BD

Sourced in United States

The FACSCanto analyzer is a flow cytometry instrument designed for high-performance cell analysis. It features advanced optics and detection capabilities for accurate and reliable data acquisition. The core function of the FACSCanto analyzer is to enable the simultaneous measurement and analysis of multiple cellular parameters in a sample.

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Lab products found in correlation

Propidium iodide, by Merck Group (4 mentions) Facsdiva software program, by BD (3 mentions) Fetal bovine serum (fbs), by Merck Group (3 mentions) Sytox green, by Thermo Fisher Scientific (3 mentions) Pe conjugated goat anti human secondary antibody, by Jackson ImmunoResearch (2 mentions) Annexin 5, by BD (2 mentions) Pac 1, by BD (2 mentions) Anti human cd63, by BD (2 mentions) Anti human cd62p, by BD (2 mentions) Anti human cd41b, by BD (2 mentions) Fc receptor block, by Miltenyi Biotec (2 mentions) Penicillin streptomycin, by Corning (2 mentions) Fluostar omega, by BMG Labtech (2 mentions) Rpmi 1640, by Corning (2 mentions) Rnase a, by Qiagen (2 mentions) Celltiter glo luminescent cell viability assay, by Promega (2 mentions) Ionomycin, by Merck Group (2 mentions) Facsdiva software, by BD (2 mentions) Alexa fluor 647 nhs ester, by Thermo Fisher Scientific (2 mentions) αpd 1, by Santa Cruz Biotechnology (2 mentions)

Spelling variants of this product

FACSCanto (2869 citations) BD FACSCanto™ II Cell Analyzer (169 citations) BD FACSCanto II analyzer (120 citations) FACSCanto Cell Analyzer (29 citations) FACSCanto II benchtop analyzer (4 citations) BD FACSCanto II Flow Cytometry Analyzer Systems (4 citations) BD FACSCanto RUO-ORANGE analyzer (3 citations) FACSCanto RUO cell analyzer (3 citations) BD Biosciences FACSCanto II Analyzer (2 citations)

66 protocols using facscanto analyzer

PD-1/PD-L1 Interaction Inhibition Assay

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A PD-L1-Fc was used to compete surface PD-1 binding of αPD-1-ABD-PE. The PD-L1-Fc is a fusion protein between mouse PD-L1 and human Fc, Sino Biological, 50010-M03H). For concurrent incubation, the PD-L1-Fc was added to a final concentration of 500 nM with Alexa Fluor 647-labeled αPD-1-ABD-PE (100 nM final concentration) into 0.5 million cells and incubated for 30 min at 4 °C or 37 °C. Then, unbound proteins were washed away by centrifugation (350 g for 5 minutes) in FACS buffer. The cells were analyzed by flow cytometry with a BD FACSCanto Analyzer (BD Biosciences). Human IgG at 500 nM was used as the control for PD-L1-Fc fusion.
In the experiment surface-bound αPD-1-ABD-PE is need to be stripped, Alexa Fluor 647-labeled αPD-1-ABD-PE was first incubated with 0.5 million of cells for 30 min at 37 °C, then PD-L1-Fc was added to a final concentration of 500 nM. The incubation mixtures were kept at 4 °C for additional 30 min before unbound proteins were washed away by centrifugation (350 g for 5 minutes) in FACS buffer. The cells were analyzed by flow cytometry with a BD FACSCanto Analyzer (BD Biosciences).

Zhao P., Wang P., Dong S., Zhou Z., Cao Y., Yagita H., He X., Zheng S.G., Fisher S.J., Fujinami R.S, & Chen M. (2019). Depletion of PD-1-positive cells ameliorates autoimmune disease. Nature biomedical engineering, 3(4), 292-305.

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PD-1/PD-L1 Interaction Inhibition Assay

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Detection of AC6 and Gαs Expression

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HEK293T cells stably expressing AC6–FLAG, AC6 (C1004A)–FLAG, and/or Gαs–HA were used to detect expression using a BD FACS canto analyzer. Briefly, 100,000 cells were washed using ice‐cold fluorescence activated cell sorting (FACS) buffer (0.5% bovine serum albumin in 1× phosphate‐buffered saline [PBS]), followed by fixation with 4% paraformaldehyde. Cells were then permeabilized with 0.2% saponin in FACS buffer and incubated with mouse monoclonal APC conjugated anti‐FLAG antibody (1:300 dilution) and/or rabbit monoclonal Alexa Fluor 488‐anti‐HA antibody (1:300 dilution) for 1 h on ice. HEK293T with APC and Alexa Fluor 488 conjugated IgG antibody were used as negative control. The cells were then washed thrice with FACS buffer and were resuspended in 200 μl of the same. The fluorescent intensity was measured using a BD FACS canto analyzer and quadrant gating was used to identify the percentage of cells expressing AC6–FLAG and/or Gαs–HA.

Bhagirath A.Y., Bhatia V., Medapati M.R., Singh N., Hinton M., Chelikani P, & Dakshinamurti S. (2021). Critical cysteines in the functional interaction of adenylyl cyclase isoform 6 with Gαs. FASEB BioAdvances, 4(3), 180-196.

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Retroviral Supernatant Generation Protocol

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To generate retroviral supernatant, GP2-293 cells were transiently transfected with either pMSCV-HrasV12-GFP or pMSCV-SV40 LTg-RFP combined with the replication-incompetent helper vector pVSV-G in a 60 mm culture dish by using Fugene 6. The cells were fed at 24 h post-transfection, and retroviral supernatant was pooled from the collection at 48 and 72 h post-transfection. The supernatant was aliquoted after centrifugation and stored at –80°C for future use. The viral titer was verified by FACS analysis (FACSCanto Analyzer, BD Biosciences, San Jose, CA) after infection to 1x 10⁵ NIH-3T3 (ATCC) mouse fibroblast cells.

Cho S., Park H., Jarboe E.A., Peterson C.M., Bae Y.H, & Janát-Amsbury M.M. (2015). Design and Characterization of Bioengineered Cancer-Like Stem Cells. PLoS ONE, 10(10), e0141172.

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Flow Cytometry Antibody Staining and Cell Sorting

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Antibodies were purchased from Biolegend or Thermo. Antibodies for flow cytometry analysis included anti-B220 (RA3–6B2), anti-NK1.1 (PK136), anti-CD11b (M1/70), anti-CD3 (2C11), anti-CD45.2 (104), and anti-Ki67 (16A8). Mouse CD1d tetramers PBS57 were obtained from the NIH tetramer core (29 (link)). Intracellular staining of Ki67 was performed using the Foxp3 Fix/perm Kit (Thermo) according to the manufacturer’s instructions. EdU was detected using the Click-iT Plus EdU Flow Cytometry Assay Kit (Thermo) following the manufacturer’s instructions. Flow cytometric analysis was performed using a FACSCanto analyzer (BD). Cell sorting was performed using a FACSAria II sorter (BD).

Zhang Y., Fung I.T., Sankar P., Chen X., Robison L.S., Ye L., D’Souza S.S., Salinero A.E., Kuentzel M.L., Chittur S.V., Zhang W., Zuloaga K.L, & Yang Q. (2020). Depletion of NK cells improves cognitive function in the Alzheimer’s Disease mouse model. Journal of immunology (Baltimore, Md. : 1950), 205(2), 502-510.

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FACS-based Cell Surface Expression Assay

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FACS-based cell surface expression assay was carried out as described previously [33 (link)]. It is to be noted here that we have used the full-length clones of both JRFL and JRCSF in all our studies. Briefly, 293T cells were transiently transfected with pSVIII-Env plasmids expressing different forms of Env protein under the control of the HIV-1 LTR and pc-tat plasmid expressing Tat protein at the ratio of 20:1. 36–48 h post transfection, cells were harvested, washed three times with FACS buffer 1 (DMEM + 10% HIFBS) and stained with varying concentrations of monoclonal antibodies (neutralizing and non-neutralizing) for 1 h at room temperature (RT). The cells were washed three times with FACS buffer 1 and then stained with PE-conjugated goat anti-human secondary antibody (1:200 dilutions, Jackson ImmunoResearch) for 1 h at RT. The cells were again washed three times with FACS buffer 2 (PBS + 10% HIFBS) and fixed with 0.5% paraformaldehyde. The stained cells were then analyzed in a FACS Canto analyzer (BD Biosciences) and data analyzed with FlowJo software (version 10.0.6, Tree Star Inc).

Das S., Boliar S., Mitra N., Samal S., Bansal M., Koff W.C, & Chakrabarti B.K. (2016). Membrane bound modified form of clade B Env, JRCSF is suitable for immunogen design as it is efficiently cleaved and displays all the broadly neutralizing epitopes including V2 and C2 domain-dependent conformational epitopes. Retrovirology, 13, 81.

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Ewing Sarcoma ALDH Activity Assay

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The Aldefluor Assay (Stem Cell Technologies) was performed on Ewing sarcoma cells according to the manufacturer's protocol. Cells were analyzed by flow cytometry using a BD FACSCanto Analyzer and FlowJo software. Viable cells were gated based on propidium-iodide exclusion. ALDH-high cells were quantified by gating the top 2% of FITC+ empty vector control cells.

Gardiner J.D., Abegglen L.M., Huang X., Carter B.E., Schackmann E.A., Stucki M., Paxton C.N., Randall R.L., Amatruda J.F., Putnam A.R., Kovar H., Lessnick S.L, & Schiffman J.D. (2017). C/EBPβ-1 promotes transformation and chemoresistance in Ewing sarcoma cells. Oncotarget, 8(16), 26013-26026.

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EGFRvIII Nanobody Screening and Binding

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EGFRvIII Nbs were screened by phage display technology according to our previously described 35 (link),51 (link). EGFRvIII Nbs with His₆ tagged were purified using Ni-NTA affinity columns. The specific purified EGFRvIII Nbs were analyzed by SDS-PAGE. For EGFRvIII Nbs specific binding assay, EGFRvIII Nbs were incubated with corresponding target cells for 30 min, then cells were stained with APC labeled anti-his tag antibody. The binding was detected by flow cytometry on a FACS Canto analyzer (BD Biosciences, USA).

Sun S., Ding Z., Gao L., Hammock B.D., Huang X., Xu Z.P., Wang X., Cheng Q., Mo F., Shi W., Xie S., Liu A., Li H., Yang X, & Lu X. (2023). A dendritic/tumor fusion cell vaccine enhances efficacy of nanobody-based CAR-T cells against solid tumor. Theranostics, 13(14), 5099-5113.

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Comprehensive Thymocyte Immunophenotyping

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Freshly isolated or cultured thymocytes were separated into single-cell suspensions and stained for various surface markers. Abs (eBioscience, San Diego, CA, USA) used for staining were conjugated to FITC, PE, PE-Cy7, or APC and raised against: CD4 (GK1.5), CD8α (53-6.7), TCRβ (H57-597), CD69 (H1.2F3), B220 (RA3-6B2), CD3ɛ (145-2C11), TCR-Vα2 (B20.1), TCR-Vβ5 (MR9-4), TCR-Vβ6 (RR4-7), TCR-Vβ8 (KJ16), NK1.1 (PK136), TER119 (TER-119), Gr-1 (RB6-8C5), CD11b (M1/70), HY TCR (T3.70), CD25 (PC61.5), and CD44 (IM7). For cell proliferation and cell death experiments, the 5-bromo-2′-deoxyuridine flow kit (BD Biosciences, San Jose, CA, USA) and annexin V apoptosis kit (BioVision, Milpitas, CA, USA) were used to detect 5-bromo-2′-deoxyuridine incorporation and dead cells, respectively. Data were collected on a FACSCanto analyzer (BD Biosciences) and analyzed using FlowJo software (TreeStar, Ashland, OR, USA).

Ho K.C., Chiang Y.J., Lai A.C., Liao N.S., Chang Y.J., Yang-Yen H.F, & Yen J.J. (2014). CBAP promotes thymocyte negative selection by facilitating T-cell receptor proximal signaling. Cell Death & Disease, 5(11), e1518-.

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Platelet Activation Markers by Flow Cytometry

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Flow cytometry was used to measure the expression levels of CD62P (P-selectin), GPIIb/IIIa (integrin α_IIbβ₃), CD63 (lysosomal glycoprotein), and phosphatidylserine on perfused platelets.
Following blood perfusion through a flow channel with a stenotic region but with no capture region, a 5 μL aliquot of blood supernatant was incubated for 20 minutes with anti-human CD62P, PAC−1, anti-human CD63, or annexin V (BD Biosciences, San Jose, CA, USA) to label P-selectin, active GPIIb/IIIa, lysosomal glycoprotein, and phosphatidylserine, respectively. Two 5 μL blood aliquots were obtained and labeled prior to perfusion. Platelet activation in one aliquot was achieved by addition of thrombin immediately prior to labeling (c = 0.1 units/mL, EMD Millipore, Billerica, MA, USA) and other aliquot was left unstimulated to serve as positive and negative activation controls, respectively. Blood was not treated with PPACK for thrombin activated positive control. To locate platelets in flow cytometry analysis, another 5 μL blood aliquot was labeled with anti-human CD41b (BD Biosciences, San Jose, CA, USA), which binds to the GPIIb subunit of GPIIb/IIIa regardless of the activation state of the receptor. Following labeling, platelets were fixed in 1% paraformaldehyde and stored at 4 °C. Analysis of 100,000 events was conducted on a FACScanto analyzer (BD Biosciences, San Jose, CA, USA).

Rahman S, & Hlady V. (2019). Downstream Platelet Adhesion and Activation Under Highly Elevated Upstream Shear Forces. Acta biomaterialia, 91, 135-143.

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