Novocyte flow cytometer

Manufactured by Agilent Technologies

Sourced in United States, China, Sweden

The NovoCyte flow cytometer is a compact and powerful instrument designed for multiparameter cell analysis. It utilizes advanced flow cytometry technology to detect and analyze various cellular characteristics, such as size, granularity, and the expression of specific markers on the cell surface or within the cell.

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Lab products found in correlation

Novoexpress software, by Agilent Technologies (83 mentions) Novoexpress, by Agilent Technologies (14 mentions) Propidium iodide, by Merck Group (11 mentions) Novoexpress 1, by Agilent Technologies (11 mentions) Lipofectamine 2000, by Thermo Fisher Scientific (10 mentions) Fetal bovine serum (fbs), by Thermo Fisher Scientific (9 mentions) Novoexpress version 1, by Agilent Technologies (8 mentions) Acea novoexpress software, by Agilent Technologies (7 mentions) Annexin 5 fitc apoptosis detection kit, by Beyotime (7 mentions) Fitc annexin 5 apoptosis detection kit 1, by BD (7 mentions) Propidium iodide, by Thermo Fisher Scientific (7 mentions) Phosphate buffered saline (pbs), by Thermo Fisher Scientific (7 mentions) Cell cycle and apoptosis analysis kit, by Beyotime (6 mentions) Cycletest plus dna reagent kit, by BD (6 mentions) Ionomycin, by Merck Group (6 mentions) Brilliant stain buffer, by BD (5 mentions) Rpmi 1640, by Thermo Fisher Scientific (5 mentions) Annexin 5 fitc pi apoptosis detection kit, by Keygen Biotech (5 mentions) Fitc annexin 5 apoptosis detection kit, by BD (5 mentions) Perm wash buffer, by BD (5 mentions)

Spelling variants of this product

NovoCyte (328 citations) NovoCyte 2060R flow cytometer (28 citations) Novocyte cytometer (13 citations) Novocyte 3000 VYB flow cytometer (6 citations) NovoCyte Penteon Flow Cytometer System (3 citations) NovoCyte 1040 flow cytometer (3 citations) NovoCyte® 1000 Flow Cytometer (2 citations) NovoCyte Benchtop flow cytometer (2 citations) NovoCyte FACS flow cytometer (2 citations) NovoCyte 3000RYB flow cytometer (2 citations)

713 protocols using novocyte flow cytometer

Annexin V-FITC and Active Caspase 3 Assay

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Cells were stained with annexin V-fluorescein isothiocyanate (FITC) and 4′,6-diamidino-2-phenylindole (DAPI) (Sigma-Aldrich, St. Louis, MI, USA) in 100 μL of binding buffer (BioLegnd, San Diego, CA, USA) for 15 min at room temperature (RT). The cells were analyzed by flow cytometry using a NovoCyte flow cytometer (ACEA Biosciences Inc., San Diego, CA, USA) after 400 μL of binding buffer was added without washing. For active caspase 3 staining, the cells were fixed and permeabilized with Cytofix/Cytoperm buffer (eBioscience, San Diego, CA, USA) and subsequently incubated with anti- active caspase 3 Abs in Perm/Wash buffer (BioLegnd) for 30 min. Staining of the isotype control immunoglobulin (Ig) G was performed in all the experiments. The active caspase 3 expression levels were analyzed by NovoCyte flow cytometer (ACEA Biosciences Inc.).

Hwang J, & Jin J.O. (2020). Attachable Hydrogel Containing Indocyanine Green for Selective Photothermal Therapy against Melanoma. Biomolecules, 10(8), 1124.

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Annexin V-PI Apoptosis Assay Protocol

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To evaluate the presence of apoptosis/necrosis, we used annexin V-Fluos (cat. no. 11828681001 Roche) and Propidium iodide (PI) and analyzed by NovoCyte flow cytometer (ACEA biosciences, Inc. San Diego, CA, USA). Between 4 and 10 × 10⁴ cells per well were cultured and treated with vemurafenib for 72 h or transfected with siRNA 48 h prior to the 72 h vemurafenib treatment. Then the cells were collected and rinsed in PBS, pelleted, and resuspended in incubation buffer (10 mmol/l HEPES/NaOH, pH 7.4, 140 mmol/l NaCl, 5 mmol/l CaCl₂) containing 1% annexin V and 1% PI for 10 min. For immunostaining against CD13 surface expression, cells were harvested and stained with FITC conjugated CD13 antibody (product No. F083101-2, Dako Sweden AB, Stockholm, Sweden) for 30 min at 4 °C in the dark followed by analysis using NovoCyte flow cytometer (ACEA biosciences).

Azimi A., Tuominen R., Costa Svedman F., Caramuta S., Pernemalm M., Frostvik Stolt M., Kanter L., Kharaziha P., Lehtiö J., Hertzman Johansson C., Höiom V., Hansson J, & Egyhazi Brage S. (2017). Silencing FLI or targeting CD13/ANPEP lead to dephosphorylation of EPHA2, a mediator of BRAF inhibitor resistance, and induce growth arrest or apoptosis in melanoma cells. Cell Death & Disease, 8(8), e3029-.

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Cell Cycle and Mitotic Analysis

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For cell cycle profile analysis cells were fixed in 70% ethanol. After 15 min incubation on ice, cells were pelleted and resuspended in 500 µl PBS containing 50 µg/ml propidium iodide (PI) (ThermoFisher, P1304MP), 0.1 mg/ml RNase A and 0.05% Triton X-100, followed by 40 min incubation at 37 °C. Cells were then washed with PBS and analyzed using the Novocyte Flow Cytometer (ACEA Biosciences, Inc.).
For G2/M checkpoint assays, 1 × 10⁶ mES cells were seeded on p60 dishes one day before exposure to 3 or 10 Gy of IR. One or 6 h later, cells were fixed as described for cell cycle profile analysis and incubated overnight at −20 °C. Fixed cells were then permeabilized for 15 min on ice using 0.25% Triton X-100 in PBS, after which mitotic cells were stained in 100 μl PBS with 1 μl anti-phospho-H3 Ser10 (1 μg/μl, Sigma-Aldrich, 06–570) for 3 h at room temperature. Alexa-488 goat α-rabbit (1:100 in 100 μl PBS; ThermoFisher, 11034) was used as a secondary antibody. Cells were analyzed using the Novocyte Flow Cytometer (ACEA Biosciences, Inc.).

Boonen R.A., Rodrigue A., Stoepker C., Wiegant W.W., Vroling B., Sharma M., Rother M.B., Celosse N., Vreeswijk M.P., Couch F., Simard J., Devilee P., Masson J.Y, & van Attikum H. (2019). Functional analysis of genetic variants in the high-risk breast cancer susceptibility gene PALB2. Nature Communications, 10, 5296.

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Assessing Mitochondrial Function and ROS Levels

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GIST 882, 882R, T1, and T1R cells seeded in 6-well plate at a 70% confluency were treated with 1 µM of imatinib. After 48 h, the cells were harvested, washed, and stained with MitoTracker Green FM and a LIVE/DEAD Viability Kit (Thermo Fisher Scientific, Waltham, MA, USA) according to the manufacturer’s instructions. The mean Mitotracker green fluorescence intensity of the live cells was measured by a NovoCyte Flow Cytometer (ACEA Biosciences, San Diego, CA, USA), and the data were analyzed using FlowJo software (FlowJo, LLC, Ashland, OR, USA).
The intracellular ROS levels of each cell line were measured using 2′,7′-dichlorofluorescin diacetate (DCFH-DA, Sigma-Aldrich, Darmstadt, Germany). The cell pellets were collected, washed with phosphate-buffered saline (PBS) and resuspended in PBS containing 5 µM of DCFH-DA. After incubation in the dark for 30 min at 37 °C, the residual DCFH-DA was removed by washing with PBS. The ROS content was measured by a NovoCyte Flow Cytometer (ACEA Biosciences, San Diego, CA, USA) using a filter with an excitation/emission = 485/535 nm.

Huang W.K., Gao J., Chen Z., Shi H., Yuan J., Cui H.L., Yeh C.N., Bränström R., Larsson C., Li S, & Lui W.O. (2020). Heterogeneity of Metabolic Vulnerability in Imatinib-Resistant Gastrointestinal Stromal Tumor. Cells, 9(6), 1333.

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Apoptosis Assay of miR-216b-5p Transfected Cells

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Apoptosis assay was performed with NovoCyte^® Flow Cytometer (ACEA Biosciences, USA). WT and TR cells were seeded in 6-well plates at a density of 25 × 10⁴ cells/well and after 24 h the cells were transfected with miR-216b-5p mimic and scrambled control. Each experiment was performed as two biological replicates. The cells were harvested and washed with PBS 48 h posttransfection and were suspended in 1X Annexin V binding buffer (10 mM Hepes/NaOH; pH: 7.4; 140 mM NaCl, 2.5 mM CaCl₂) in a concentration of 1 × 10⁶ cells/mL. Five μL Annexin V-FITC (640906, BioLegend) and five μL PI (421301, BioLegend) (0.5 μg/μL) were added and the cells were incubated in dark for 15 min. After the incubation period, 400 μL Annexin V binding buffer was added on the cells and data was obtained in an 1 h by NovoCyte^® Flow Cytometer (ACEA Biosciences, USA).

DOLAPÇI İ.B., NOYAN S., POLAT A.Y., GÜRDAL H, & DEDEOĞLU B.G. (2023). miR-216b-5p promotes late apoptosis/necroptosis in trastuzumab-resistant SK-BR-3 cells. Turkish Journal of Biology, 47(3), 199-207.

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Flow Cytometry Analysis of EV-Induced T Cell Activation

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Jurkat 76 TPR cells (100,000 cells per well) were incubated with EVs or MoDCs at different T cell:EV or MoDC ratios (2:1, 1:1, 1:2, and 1:5) for 16 h in a 96-well flat bottom plate (Corning). Positive control cells were incubated with 50 ng/mL PMA (Merck) and 1 μg/mL ionomycin (Merck). The volume of each well was 200 μL. Following incubation, cells were washed and analyzed for GFP expression using the NovoCyte flow cytometer (ACEA Biosciences). Alternatively, Jurkat 76 TPR cells were analyzed for the CD69 activation marker. Jurkat 76 TPR cells were incubated with EVs under the same conditions, stained with anti-human CD69, washed, and analyzed using the NovoCyte flow cytometer (ACEA Biosciences).

Ishina I.A., Kurbatskaia I.N., Mamedov A.E., Shramova E.I., Deyev S.M., Nurbaeva K.S., Rubtsov Y.P., Belogurov AA J.r., Gabibov A.G, & Zakharova M.Y. (2024). Genetically engineered CD80–pMHC-harboring extracellular vesicles for antigen-specific CD4+ T-cell engagement. Frontiers in Bioengineering and Biotechnology, 11, 1341685.

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Immunophenotyping of Cells and Extracellular Vesicles

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Cells were washed with PBS and re-suspended in PBS with fluorescent antibodies. HeLa cells were stained with anti-human CD80 and anti-human HLA-DR. Jurkat 76 TPR cells were stained with anti-human CD4, anti-human CD3-APC, and anti-human TCR. The staining was performed for 30 min at 4°C. Then, cells were washed with PBS and analyzed using the NovoCyte flow cytometer (ACEA Biosciences). Centrifuging steps were performed at 300 g at 4°C.
EVs were washed with DMEM, 10% FBS, and 1% pluronic F-127 and re-suspended in the same medium with fluorescent antibodies anti-human CD80 and anti-human HLA-DR. The staining was performed for 30 min at 4°C. Then, EVs were washed with the same medium and analyzed using the NovoCyte flow cytometer (ACEA Biosciences). Centrifuging steps were performed at 5000 g at 4°C.

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Multiparametric Immunophenotyping in Cell Samples

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Cell counts and viability were determined using a NovoCyte Flow Cytometer (ACEA Biosciences, San Diego, CA, USA) and 7-aminoactinomycin D (7-AAD, Interchim, Montluçon, France). Immunophenotyping of T cells (including homing receptors), dendritic cells (DC), innate lymphoid cells (ILC), and inflammatory cells was performed using four pre-optimized antibody panels presented in the Supplementary Materials (Table S1). Data acquisition was performed on an Attune™ NxT Flow Cytometer (Thermo Scientific™, Waltham, MA, USA) or on a NovoCyte Flow Cytometer, depending on the cell subtypes, and analyses were performed using FlowJo^® (Version 10, ACEA Biosciences, San Diego, CA, USA). The gating strategies are illustrated in Figures S1–S4 in the Supplementary Materials.
The various analyses described above were carried out on all the samples at the same time.

Briard M., Guinot M., Grauso M., Guillon B., Hazebrouck S., Bernard H., Bouchaud G., Michel M.L, & Adel-Patient K. (2022). Route of Sensitization to Peanut Influences Immune Cell Recruitment at Various Mucosal Sites in Mouse: An Integrative Analysis. Nutrients, 14(4), 790.

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Cell Cycle Arrest Analysis of miR-34a Delivery

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Cell cycle arrest analysis was performed to evaluate whether LbL NP-mediated delivery of miR-34a could induce cell cycle arrest and, if so, in which phase of the cell cycle. MDA-MB-231 cells were seeded in 60-well plates at 50,000 cells/well for approximately 22 hr. Then, cells were either left untreated or treated with LbL NPs carrying miR-co or miR-34a at a dose of 250 nM miRNA in complete medium. After 24 hr of treatment, cells were washed three times with 1X PBS and replenished with fresh medium. After a total treatment of 66 hr, the cells were washed with 1X PBS, stained with DRAQ5™ (Thermo Fisher) DNA probe dye at a concentration of 15 μM in 1X PBS, and incubated at 37°C in a 5% CO₂ humidified incubator. After 30 min, the staining solution was removed and the cells trypsinized. Once the cells were non-adherent, the trypsin was neutralized with media and the cells transferred to a 15 ml centrifuge tube. The cells were centrifuged at 300g to pellet the cells; the supernatant was removed, and the cell pellet was resuspended in 300 μL of 1X PBS. Fluorescence intensity of DRAQ5™ was evaluated by flow cytometry using a NovoCyte flow cytometer (ACEA Biosciences) with 488 nm excitation and 530/30 nm detection. Cell cycle analysis was performed using a default algorithm provided by the NovoCyte flow cytometer (ACEA Biosciences).

Kapadia C.H., Ioele S.A, & Day E.S. (2019). Layer-by-layer assembled PLGA nanoparticles carrying miR-34a cargo inhibit the proliferation and cell cycle progression of triple-negative breast cancer cells. Journal of biomedical materials research. Part A, 108(3), 601-613.

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Cell Cycle and Apoptosis Analysis

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Cell cycle assays were performed using a Cell Cycle Detection kit (Beyotime). After digestion and collection, the cells were washed twice with precooled PBS and fixed with 70% ethanol (Aladdin, Shanghai, China) at 4°C overnight. Then, the ethanol was removed, and the cells were washed with PBS. After adding 100 μl RNaseA to the samples, the cells were incubated in a 37°C water bath for 30 minutes. Finally, 500 μl PBS containing 50 μg/ml propidium iodide (PI) was added to the samples and incubated at 4°C in the dark for 30 minutes. A NovoCyte^TM Flow Cytometer (ACEA Biosciences, San Diego, USA) was used to analyze the results. An Annexin V-FITC kit (Beyotime) was used to assess cell apoptosis. After collection, the cells were washed twice with PBS, and 500 μl binding buffer was added to resuspend the cells. The samples were mixed well with 5 μl Annexin V-FITC and 10 μl PI and incubated at room temperature in the dark for 15 minutes. A NovoCyte^TM Flow Cytometer (ACEA Biosciences) was used to analyze the results.

Lai T., Qiu H., Si L., Zhen Y., Chu D, & Guo R. (2022). Long noncoding RNA BMPR1B-AS1 facilitates endometrial cancer cell proliferation and metastasis by sponging miR-7-2-3p to modulate the DCLK1/Akt/NF-κB pathway. Cell Cycle, 21(15), 1599-1618.

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