Accuri c6 ow cytometer

Manufactured by BD

Sourced in United States

The BD Accuri C6 is a flow cytometer designed for analytical applications. It is capable of measuring various parameters of cells or particles in a sample, including size, granularity, and fluorescence. The device uses laser technology to detect and analyze the properties of individual cells or particles as they pass through a fluid stream.

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BD Accuri C6 Plus ow cytometer Accuri C6 ow cytometer and software BD Accuri ow cytometer C6 ow cytometer BD Accuri C6 cytometer BD Accuri C6 Ow cytometer BD Accuri cytometer BD Accuri

26 protocols using accuri c6 ow cytometer

Cell Cycle Analysis and Apoptosis Assay

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Cells were collected with 0.25% trypsin and washed twice with cold PBS, then xed with 70%-75% frozen ethanol at -20 °C for 1 h or xed overnight at 4 °C. Subsequently, cells were washed once with cold PBS and then resuspended in 200-500 µl cold PBS. Then, 20 µl RNase A solution was added, and samples were incubated in a 37 °C water bath for 30 min. Finally, 400 µl PI stain solution (BestBio, Shanghai, China) was mixed gently and incubated for 30-60 at 4 °C in the dark. The results were detected with an Accuri C6 ow cytometer (Becton Dickinson, Franklin Lakes, New Jersey, USA).
Apoptosis analyses NCI-H446 and H1299 cells were plated on six-well plates at a concentration of 1 × 10 5 cells/well.
Transfection experiments were performed the following day. Cells were harvested after 48 h and washed twice with PBS. Cells were resuspended in 400 µl 1× buffer, then stained with 5 µl Annexin-V-FITC and 5 µl PI (50 μg/ml) with an Annexin V FITC Apop Dtec Kit (BD Biosciences, San Jose, CA, USA) in the dark for 15 min at room temperature, and detected with an Accuri C6 ow cytometer (Becton Dickinson, Franklin Lakes, New Jersey, USA).

Jia S., Wang L., Jiao Y., Guo B., Wang L., Peng L., & Zhang L. (2021). Ssrp1 Promotes Lung Cancer Progression By Blocking The Wnt Pathway and Is Negatively Regulated By Mirna-28-5p.

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Sperm DNA Fragmentation Analysis

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Evanson's method is considered to be the gold standard for DFI detection and has been widely used worldwide (Alharbi, Hamouche, Phillips, Kadoch, & Zini, 2019) (link). In our study, the human sperm nuclear integrity staining kit (Xingbo Biological, Shanghai, China) was used to detect DFI (Du et al., 2017) . The chromatin structure of damaged sperm is loose; thus, the DNA denatures to form a single strand under the action of acid while the DNA of normal sperm maintains the double-stranded structure. Acridine orange (AO) emits green uorescence on combination with double-stranded DNA and yellow or red uorescence on combination with single stranded DNA. Sperms were stained with AO and analyzed using ow cytometry. A certain amount of semen was diluted to 100 μL with solution A to derive a nal concentration of (1-2) ×10 6 sperm cells/mL. After adding 200 μL of solution B (cells were placed on ice for operation), 600 μL of solution C was added after a certain time interval for 30 s. At least 5,000 sperm cells from each sample were analyzed using BD Accuri® C6 ow cytometer (BD Bioscience, Shanghai, China), and the results were analyzed using Accuri® CFlow® software (BD Bioscience, Shanghai, China).

Gu B., Wang S., Liu F., Song Y., Li J., Ni Y., Chen M., Hu J., Ouzhu L., Li Z., Liu L., Li X, & Liu X. (2022). Same total normal forms sperm counts of males from Lhasa and Shanghai, China. Environmental science and pollution research international, 29(13).

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Apoptosis Evaluation of OA and MA

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Evaluation of the apoptotic effect of OA and MA on MCF7 cell lines through the phosphatidylserine (PS) in the cell membrane was performed according to the manual of the FITC Annexin V Apoptosis Detection Kit (Cat. No.: 556547, BD Pharmingen, USA) protocol by counting 3.5 × 105 cells on the BD ACCURI C6 ow cytometer (BD Biosciences Pharmingen). After OA and MA were added to the cells at the desired dosage, their apoptotic effect was examined for 72 hours. The group that was not treated with OA and MA was considered as the control group.

Avci Ç.B., Söğütlü F., Özateş N.P., Shademan B., & Gündüz C. (2021). Enhanced Anti-Cancer Potency Using A Combination of Oleanolic Acid and Maslinic Acid to Control Treatment Resistance in Breast Cancer.

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Evaluating Cell Proliferation and Cell Cycle

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CCK-8 (EZBioscience, Roseville, MN) assay was used to evaluate cell proliferation. Approximately 10 4 cells were seeded in 96-well plates. After they adhered to the wall, they were transfected with OE-circKANSL1L, OE-NC, si-circKANSL1L, or si-NC. Six hours after transfection was recorded as 0 h. CCK-8 was added at 0, 12, 24, and 36 h, followed by incubation for 1 h. Absorbance was then measured at 450 nm using a microplate reader (Thermo Fisher Scienti c, Waltham, MA).
Flow Cytometric Cell Cycle Analysis C 2 C 12 cells were transfected with OE-circKANSL1L, OE-NC, si-circKANSL1L, and si-NC. After 48 h, the cells were collected, xed with 75% ethanol, and stored overnight at -20°C. They were then resuspended in 500 μL PI/RNase staining buffer solution (BD Biosciences, Franklin Lakes, NJ) and incubated at 37°C for 30 min. A BD Accuri C6 ow cytometer and FACSDiVa software (BD Biosciences) were used to perform ow cytometric analysis.

Zhuang X., Lin Z., Luo J., Chen T., Xi Q., Zhang Y., Sun J., & Xie F. (2021). Identification of CircRNA-Associated ceRNA Networks Using Longissimus Thoracis of Pigs of Different Breeds and Growth Stages.

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Apoptosis Analysis by Flow Cytometry

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For the apoptosis analysis, 1 × 10 6 cells were seeded in a 60 mm plate, treated with OE-CircCCT3 or CCT3 siRNA, and cultured for 24 h. After harvesting, 1 × 10 5 cells were suspended in a 1x binding buffer provided with the Annexin V-FITC Apoptosis Detection kit II (BD Bioscience, San Jose, CA, USA), then stained with FITC Annexin V(BD Bioscience) and PI (Sigma-Aldrich, St. Louis, MO, USA). Fluorescence was detected with a BD Accuri C6 ow cytometer (BD Bioscience), and the data were analyzed with the BD Accuri C6 software (BD Bioscience).

Hou J., Men X., Yang L., Han E., Chunqi H., & Liu L. (2021). CircRNA CircCCT3 Acts as a Sponge of miR-613 to Promote Tumor Growth of Pancreatic Cancer Through Regulating VEGFA/VEGFR2 Signaling.

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Quantifying CD36 Cell Surface Expression

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To determine cell surface expression of CD36, cells were labelled with anti-human CD36 antibody (FA6-152, Stem Cell Technologies, Vancouver, Canada) or isotype control (IgG 1 κ) at a concentration of 10 µg/ml in RPMI1640 media (Sigma-Aldrich, Merck, Sydney Australia) containing 10% FBS. Cells were then washed and stained with a secondary antibody, goat anti-mouse IgG H&L Dylight 650 (Clone ab96882, Abcam, Cambridge, United Kingdom) at a concentration of 5 µg/ml. Cell viability was determined via 7AAD (BD Biosciences) staining at a 1:20 dilution. Samples were processed using a BD Accuri C6 ow cytometer (BD, Becton, Dickinson and Company, New Jersey, USA) and data analyzed using FCS Express 4 Flow Research Edition (De Novo software, Los Angeles, USA).
CD36 knockdown using siRNA CD36 targeting small interfering RNAs (siRNAs) Trilencer-27 Human siRNA duplexes at 20µM were purchased (Origene, Maryland, USA) with three CD36 targeting siRNAs (A, B and C) plus a non-targeting scrambled (SCR) siRNA used as a negative control. Transfection was performed using Lipofectamine RNAiMAX (Invitrogen by Life Technologies, California, USA) as per the manufacturer's instructions with the knockdown maximized at 5nM and validated by ow cytometry.

Martini C., DeNichilo M., King D.P., Cockshell M.P., Ebert B., Dale B., Ebert L.M., Woods A.E., & Bonder C.S. (2021). CD36 Promotes Vasculogenic Mimicry in Melanoma by Mediating Adhesion to the Extracellular Matrix.

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AuNPs Uptake by Macrophages

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The AuNPs exposed J774.1A Ms were incubated with 1 µm-diameter FluoSpheres® Carboxylate-Modi ed Microspheres (1 µm, ThermoFisher, cat. no.: F8851), at a ratio of 10 microspheres per cell for 6 h at 37 °C in a 5% CO 2 incubator. The cells were analysed for their uorescence on a BD Accuri™ C6 ow cytometer (BD Biosciences) and FCS Express V6 (De Novo Software).

Dey A.K., Gonon A., Pécheur È., Pezet M., Villiers C., & Marche P.N. (2020). Impact of Gold Nanoparticles on the Functions of Macrophages and Dendritic Cells.

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Apoptosis Assay of BMSCs

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STS (500 nM for working solution) induced BMSCs apoptosis for 2 h, 4 h, 6 h, and 12 h. 5 µl Annexin V and 10 µl PI were stained with apoptotic BMSCs for 30 min, and the apoptotic ratio was detected by Flow cytometry (BD Accuri C6 ow cytometer). Cells positive for FITC Annexin V and negative for PI (Q3) are undergoing apoptosis. Necrotic cells stain negative for FITC Annexin V and positive for PI (Q1). Cells positive both for FITC Annexin V and PI (Q2) are either in the end stage of apoptosis or dead. Alive cells are negative for both FITC Annexin V and PI (Q4).

Li M., Xing X., Huang H., Cheng L., Gao X., Tang Q., Xu X., Sun Y., Liao L., & Tian W. (2021). The Role of Apoptotic Bodies From Different Stages of Apoptosis in Maintaining the Vitality of BMSCs.

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Radiation-Induced Apoptosis Analysis

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Cells were treated with 8 Gy radiation for 72 h, then stained with FITC-Annexin V and propidium iodide (PI) using an FITC Annexin V Apoptosis Detection Kit (BD Bioscience) following the manufacturer's instructions. Apoptosis was measured using a BD Accuri C6 ow cytometer.

Zhao M., Wang Y., Zhao Y., He S., Zhao R., Song Y., Cheng J., Gong Y., Xie J., Wang Y., Hu B., Tian L., & Huang Q. (2020). Caspase-3 Silencing Attenuates Tumor Repopulation Via Impairing Radiation-Induced DNA Damage Response and Downstream Pathway in Non-Small-Cell Lung Cancer.

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Apoptosis Assessment by Flow Cytometry

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The different groups of cells were stained with FITC-Annexin V and propidium iodide (PI) in the dark with the Annexin V-FITC/PI Apoptosis Detection Kit (BD556547, BD Bioscience, USA) according to the manufacturer's instructions. Then, each sample was washed and analyzed with an Accuri™ C6 ow cytometer (BD Bioscience, USA).

Chen P., Chen X., Zhang H., Chen J., Lin M., Qian H., Gao F., Chen Y., Gong C., Zheng X, & Zheng T. (2023). Dexmedetomidine Regulates Autophagy via the AMPK/mTOR Pathway to Improve SH-SY5Y-APP Cell Damage Induced by High Glucose. Neuromolecular medicine, 25(3).

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