Cmfda dye

Manufactured by Thermo Fisher Scientific

Sourced in United Kingdom, Ireland

CMFDA dye is a fluorescent probe used for live-cell imaging applications. It is a cell-permeant dye that becomes fluorescent upon reacting with thiols present in the cell, enabling the visualization and tracking of live cells.

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Lab products found in correlation

Collagen type 1, by Thermo Fisher Scientific (1 mentions) Matrigel, by BD (1 mentions) Cmac dye, by Thermo Fisher Scientific (1 mentions) Deepred dye, by Thermo Fisher Scientific (1 mentions) Cm dil, by Thermo Fisher Scientific (1 mentions) X cite, by Excelitas (1 mentions) Metamorph software, by Molecular Devices (1 mentions)

Close spelling variants of this product

CellTracker™Green CMFDA fluorescent dye CellTracker Green CMFDA Dye CMFDA Cell Tracker dye CellTracker CMFDA dye Green CMFDA Dye CMFDA) green fluorescent dye Molecular Probes CellTracker Green CMFDA Dye CMFDA

7 protocols using cmfda dye

Evaluating Chemotherapy Resistance in Bone Marrow Cells

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Different BM cells were labeled with CMFDA dye (Invitrogen) for 2 min at room temperature in the dark and washed 3 times with 10 mL of PBS supplemented with 0.5% FBS to remove unbound dye. CMFDA-labeled BM cell populations were cultured overnight in direct contact with MM cells followed by 24h treatment with chemotherapeutics or vehicle control. Mouse cells were treated with 0.25–1 μM doxorubicin or 7.5μM melphalan; human cells were treated with 0.5–2 μM doxorubicin or 5–10μM melphalan. Cells were then collected and apoptosis was evaluated by flow cytometry in the gated population of MM CMFDA-negative cells. Specific chemotherapy-induced apoptosis was calculated by subtracting background values (apoptosis of cells treated with vehicle control) from values of apoptosis in cells undergoing chemotherapy treatment. Values of specific chemotherapy-induced apoptosis are presented in figures.

Ramachandran I.R., Condamine T., Lin C., Herlihy S.E., Garfall A., Vogl D.T., Gabrilovich D.I, & Nefedova Y. (2015). Bone marrow PMN-MDSCs and neutrophils are functionally similar in protection of multiple myeloma from chemotherapy. Cancer letters, 371(1), 117-124.

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Intestinal Tissue-on-a-Chip with Immune Cells

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Both chambers of the chip were coated with type I collagen (30 μg/ml; Gibco) and Matrigel (100 μg/ml; BD Biosciences, Bedford, MA, United States) in serum-free MEM, which was injected into the microchannels and incubated for 1 h before cell plating. Caco-2 cells (1 × 10⁵ cell/cm²) were stained with 5 mM green cell-tracker (CMFDA Dye, Invitrogen) were seeded in the lower microculture chamber, and the chip was turned upside down and incubated at 37°C allowing the seeded intestinal epithelial cells to grow on the membrane surface, Caco-2 static culture for 1 day, then continuously perfused into Caco-2 chambers at 60 μl/h using a multi-channel injection pump (LSP04-1A, LONGER Halma, England). After 2 days, HUVECs (1 × 10⁵ cells/cm²) stained with 5 mM red cell-tracker (CMPTX Dye, Invitrogen) were seeded into the upper microculture chamber and static culture for 1 day, then continuously perfused into all chambers at 60 μl/h using a multi-channel injection pump. After 3 days, PBMC (1 × 10⁷ cells/ml) with fresh antibiotic-free culture medium (50% HUVECs culture medium and 50% PBMC medium) was continuously perfused into the upper chamber (capillary side) at 60 μl/h using a multi-channel injection pump.

Zhao W., Yao Y., Zhang T., Lu H., Zhang X., Zhao L., Chen X., Zhu J., Sui G, & Zhao W. (2022). Primary exploration of host–microorganism interaction and enteritis treatment with an embedded membrane microfluidic chip of the human intestinal–vascular microsystem. Frontiers in Bioengineering and Biotechnology, 10, 1035647.

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Intestinal Epithelial-Vascular Coculture

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Before the microdevice was assembled, all tubes and PDMS plates were pre-treated with 75% ethanol for 12 h, and the entire system was then dried in oven at 60°C. The dried devices were exposed to ultraviolet light for 30 min. The porous membrane was coated with collagen type I hydrogel at 37°C. The Caco-2 cells (1 × 10⁵ cells/cm²) stained by blue cell-tracker (CMAC Dye, Invitrogen) were seeded on the porous PDMS membrane and incubated at 37°C for 3–4 h, allowing the seeded intestinal epithelial cells to attach to the membrane surface. Then, HUVECs (1 × 10⁵ cells/cm²), which were stained by green cell-tracker (CMFDA Dye, Invitrogen) and wrapped by type I collagen gel, were seeded onto the basal of the membrane. After vascular endothelial cell attached to the membrane, the microdevice was assembled and ran as described above.

Jing B., Wang Z.A., Zhang C., Deng Q., Wei J., Luo Y., Zhang X., Li J, & Du Y. (2020). Establishment and Application of Peristaltic Human Gut-Vessel Microsystem for Studying Host–Microbial Interaction. Frontiers in Bioengineering and Biotechnology, 8, 272.

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Melanoma Cell Cytotoxicity Assay

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Melanoma cells were cultured in 96 well plates and then exposed to 2.5 µM, 5 µM, 10 µM of BMS202-NDs or to 10 µM of BMS202 alone for 6 h and then co-cultured with 100,000 hPBMCs for an additional 24 h. Then hPBMCs/melanoma cell lines were washed, fixed in 3% paraformaldehyde (PFA), and then stained with cytotoxic T cell marker (anti-CD8 antibodies), whereas melanoma cell lines were probed with cell tracker (green—C7025, CMFDA Dye, ThermoFisher Scientific, Dublin, Ireland).

Alharbi B., Qanash H., Binsaleh N.K., Alharthi S., Elasbali A.M., Gharekhan C.H., Mahmoud M., Lioudakis E., O’Leary J.J., Doherty D.G., Mohamed B.M, & Gray S.G. (2023). Proof of concept nanotechnological approach to in vitro targeting of malignant melanoma for enhanced immune checkpoint inhibition. Scientific Reports, 13, 7462.

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Assessing Cell-Cell Interactions in Vitro

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IFN-γ activated THP-1 cells were labelled with viability compatible CMFDA dye (Cat#C7025, Thermo Scientific) at 5 μM and then loaded with Staphylococcal Enterotoxin B (SEB) (Item# BT202, Toxin Technology) at 5 μg/mL for 30 min. CD4+ T cells were labelled with 0.5 μM viability compatible Deep red dye (Cat# C34565, Thermo Scientific) and then exposed to ManLAM as described above in RPMI 1640 media. Each cell type was washed and resuspended in M10 at 50,000 cells/μL. Resuspended cells were co-cultured for 30 min at 37°C in 5% CO₂ by adding 10 μL of each cell type in a 96-well U-bottom plate. Cells then were fixed in 3% PFA, 3% sucrose in 1xPBS before acquisition on an LSR Fortessa flow cytometer. At least 50,000 events were acquired per condition.

Mwebaza I., Shaw R., Li Q., Fletcher S., Achkar J.M., Harding C.V., Carpenter S.M, & Boom W.H. (2023). Impact of Mycobacterium tuberculosis Glycolipids on the CD4+ T Cell–Macrophage Immunological Synapse. The Journal of Immunology Author Choice, 211(9), 1385-1396.

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Real-Time T-cell Tracking Assay

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After transfection for 24 h, the treated cells were seeded on a glass bottom, light-resistant 24-well plate for 12 h, subjected to staining with the live-cell fluorescent dye CM-Dil (Thermo Fisher), and co-cultured with activated human primary T cells or Jurkat cells labeled with the CMFDA dye (Thermo Fisher) at a ratio of 1:10. A high-content screening system with Harmony software, version 4.9 (PerkinElmer, USA) or a confocal microscope with Volocity software, version 6.1.1 (PerkinElmer) was used to track the T-cell fluorescence status in real-time. Green fluorescence was observed in surviving T cells, and all living and apoptotic cells could be observed in bright fields.

Wang J., Ge J., Wang Y., Xiong F., Guo J., Jiang X., Zhang L., Deng X., Gong Z., Zhang S., Yan Q., He Y., Li X., Shi L., Guo C., Wang F., Li Z., Zhou M., Xiang B., Li Y., Xiong W, & Zeng Z. (2022). EBV miRNAs BART11 and BART17-3p promote immune escape through the enhancer-mediated transcription of PD-L1. Nature Communications, 13, 866.

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Microfluidic Cell Lysis Imaging

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GBM TICs were first labeled with 5 μM CMFDA dye (Thermo Fisher Scientific) per the manufacturer’s instructions. Cells were then seeded into a microwell at the terminus of the enclosed SCAMPR devices and allowed to adhere for at least 1 hour. Next, the PDMS lid was removed and the device was either left in PBS (Open condition) or layered with a 100 μm thick 5% w/v agarose lid (Agarose Lid condition) (Life Technologies). The device was then placed in a custom lysis chamber with an optically transparent bottom to allow for imaging. Lysis buffer was poured over the device and images were taken every 0.5 seconds for 60 seconds. Time-lapse imaging was taken using MetaMorph software (Molecular Devices) with a 200 ms exposure time, at 1 × 1 pixel binning through a 10x magnification objective (Olympus UPlan FLN, NA 045) on an Olympus IX71 inverted fluorescence microscope equipped with an camera (Photometrics Coolsnap HQ2), motorized stage (Applied Scientific Instrumentation), FITC/GFP filter cube (Omega XF100-3, Ex/Em: 445–495/505–575nm), and shuttered mercury lamp light source (X-cite, Lumen Dynamics).

Lin J.M., Kang C.C., Zhou Y., Huang H., Herr A.E, & Kumar S. (2018). Linking Invasive Motility to Protein Expression in Single Tumor Cells. Lab on a chip, 18(2), 371-384.

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