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Cmf2hc

Manufactured by Thermo Fisher Scientific
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The CMF2HC is a compact, high-performance centrifuge designed for a variety of laboratory applications. It features a user-friendly interface, programmable speed and time settings, and a brushless motor for quiet operation. The centrifuge can accommodate a range of rotor options to meet the needs of diverse sample types and volumes.

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Lab products found in correlation

H2dcfda, by Thermo Fisher Scientific (10 mentions) Celltracker blue, by Thermo Fisher Scientific (4 mentions) Cm h2dcfda, by Thermo Fisher Scientific (3 mentions) Magic red cathepsin b assay kit, by ImmunoChemistry Technologies (3 mentions) Ti2eu, by Nikon (2 mentions) Cd11c microbeads, by Miltenyi Biotec (2 mentions) Tamra, by Thermo Fisher Scientific (2 mentions) Cmf2hc, by Beyotime (1 mentions) H2dcfda, by Beyotime (1 mentions) Celltracker blue dye, by Thermo Fisher Scientific (1 mentions) Dcfh da, by Beyotime (1 mentions) Dmi4000b, by Leica (1 mentions) Gsh assay kit, by Beyotime (1 mentions) Celltrace violet, by Thermo Fisher Scientific (1 mentions) Carboxyfluorescein succinimidyl ester (cfse), by Thermo Fisher Scientific (1 mentions) Anti cd4, by Thermo Fisher Scientific (1 mentions) Anti cd11c, by Thermo Fisher Scientific (1 mentions) Dnase 1, by Roche (1 mentions) Anti il 2, by Thermo Fisher Scientific (1 mentions) Anti foxp3, by BD (1 mentions)

Close spelling variants of this product

CellTracker Blue CMF2HC (30 citations) CellTracker™ Blue CMF2HC Dye (7 citations) 4-chloromethyl-6,8-difluoro-7-hydroxycoumarin (CMF2HC, (4 citations) Cell Tracker Blue CMF2HC Molecular Probes (3 citations) CellTracker Blue 4-chloromethyl-6,8-difluoro-7-hydroxycoumarin (CMF2HC; (3 citations) CMF2HC) Cell Tracker Blue dye (2 citations)

26 protocols using cmf2hc

1

Measuring Oxidative Stress in MII Oocytes

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After 42–44 h, ROS and GSH levels of MII oocytes were detected through H2DCFDA (Beyotime, Shanghai, China) and CMF2HC (Beyotime, Shanghai, China). Each group was stained by H2DCFDA and CMF2HC for 10 μM and 30 min at room temperature, then 4 μL PBS droplets were placed in a 35 mm small dish (Thermo Fisher Scientific, Wilmington, DE, USA), and each group of oocytes was transferred to different dishes in turn. Stained samples were imaged with an inverted fluorescence microscope (Ti2eU; Nikon, Tokyo, Japan) using ImageJ software (version 8.0.2, NIH, Bethesda, MD, USA) to examine fluorescence intensity. For each experimental group, 30 to 60 oocytes were selected for analysis.
Wang C.R., Yuan X.W., Ji H.W., Xu Y.N., Li Y.H, & Kim N.H. (2024). Chrysoeriol Improves the Early Development Potential of Porcine Oocytes by Maintaining Lipid Homeostasis and Improving Mitochondrial Function. Antioxidants, 13(1), 122.
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2

Oocyte ROS and GSH Quantification

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To measure ROS levels, oocytes were incubated with 10 μM 2′,7′-dichlorodihydrofluorescein diacetate (H2DCFDA; Thermo Fisher Scientific, Waltham, USA) for 30 min, followed by spectroscopic
analysis (green fluorescence, UV filters, 490 nm). To measure GSH levels, oocytes were incubated with 10 μM Cell Tracker Blue dye 4-chloromethyl-6,8-difluoro-7-hydroxycoumarin (CMF2HC)
(Thermo Fisher Scientific) for 30 min, followed by spectroscopic analysis (blue fluorescence, UV filters, 370 nm). The fluorescence intensity of the oocytes was analyzed using ImageJ
software.
YAO X., JIANG H., LIANG S., SHEN X., GAO Q., XU Y.N, & KIM N.H. (2018). Laminarin enhances the quality of aged pig oocytes by reducing oxidative stress. The Journal of Reproduction and Development, 64(6), 489-494.
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3

Intracellular ROS and GSH Levels in Porcine Embryos

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To determine intracellular ROS abundance, porcine 4-cell- or blastocyst-stage embryos were incubated with 10 µM 2′,7′-dichlorodihydrofluorescein diacetate (DCFH-DA; S0033S, Beyotime, Shanghai, China) for 15 min. Furthermore, porcine embryos were cultured in IVC medium with or without 1 µM MA for 2 or 6 days under oxidative damage conditions (200 µM H2O2 preincubation for 30 min) to define whether MA reversed the oxidative damage in porcine embryos. To determine intracellular GSH levels, porcine 4-cell- or blastocyst-stage embryos were incubated with 10 μM 4-chloromethyl-6,8-difluoro-7-hydroxycoumarin (CMF2HC; C12881, Thermo Fisher, Shanghai, China) for 30 min. The fluorescence signals of both ROS and GSH were captured in tagged image file format (TIFF) using a digital camera connected to the fluorescence microscope, and fluorescence intensities were analysed using NIH ImageJ software.
Yang T.T., Qi J.J., Sun B.X., Qu H.X., Wei H.K., Sun H., Jiang H., Zhang J.B, & Liang S. (2023). Maslinic Acid Supplementation during the In Vitro Culture Period Ameliorates Early Embryonic Development of Porcine Embryos by Regulating Oxidative Stress. Animals : an Open Access Journal from MDPI, 13(6), 1041.
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4

Intracellular ROS and GSH Quantification

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Intracellular ROS and GSH levels were measured with CM-H2DCFDA (Invitrogen, Carlsbad, CA, USA) and CMF2HC (Invitrogen), respectively. D2 embryos and D6 blastocysts were incubated for 30 min in DPBS/PVA containing 5 μmol/L CM-H2DCFDA or 10 μmol/L CMF2HC. After incubation, embryos and blastocysts were washed in DPBS/PVA, and fluorescence was observed using a fluorescence microscope (DMI 4000B; Leica) with ultraviolet filters (460 nm for ROS and 370 nm for GSH). The fluorescence signal intensities were analyzed using ImageJ software (version 1.47; National Institutes of Health, Bethesda, MD, USA) after normalization through subtraction of the background intensity to that of control embryos.
Joo Y.E., Jeong P.S., Lee S., Jeon S.B., Gwon M.A., Kim M.J., Kang H.G., Song B.S., Kim S.U., Cho S.K, & Sim B.W. (2023). Anethole improves the developmental competence of porcine embryos by reducing oxidative stress via the sonic hedgehog signaling pathway. Journal of Animal Science and Biotechnology, 14, 32.
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5

Quantifying Intracellular ROS and GSH

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To detect the ROS and GSH levels, MII stage oocytes were sampled in medium with added P4 (100 µM) or RU-486 (25 µM) for determination of their intracellular ROS and GSH levels. For detection of the ROS levels, the oocytes were incubated with 10 µM H2DCFDA for 15 min (green fluorescence, UV filters, 460 nm). For detection of the GSH levels, the oocytes were incubated with 10 µM CMF2HC (Invitrogen) for 15 min (blue fluorescence, UV filters, 370 nm). The same procedures were followed for all groups of oocytes, including incubation, rinse, mounting and imaging. Image J software was used to analyze the fluorescence intensities of the oocytes. Three independent experiments were performed.
Yuan B., Liang S., Jin Y.X., Kwon J.W., Zhang J.B, & Kim N.H. (2016). Progesterone influences cytoplasmic maturation in porcine oocytes developing in vitro. PeerJ, 4, e2454.
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6

Quantifying Oxidative Stress in Oocytes

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To determine the levels of intracellular ROS and GSH in living oocytes, oocytes were stained for 30 min in PVA-PBS containing 10 μM 2´,7´-dichlorodihydrofluorescein diacetate (H2DCFDA; Invitrogen, Carlsbad, CA) or 4-chloromethyl-6,8-difluoro-7-hydroxycoumarin (CMF2HC; Invitrogen), respectively, at 37°C. After incubation, oocytes were washed three times and then transferred into PVA-PBS droplets and covered with mineral oil. Fluorescence signals were observed using a fluorescence microscope (DMi8). ROS and GSH levels were quantified by determining the fluorescence intensity in the cytoplasm of the oocytes using ImageJ. For measuring cathepsin B activity, oocytes were stained according to the Magic Red Cathepsin B Assay Kit (Immunochemistry Technologies LLC, Bloomington, MN) protocol. The same procedure was used for all replicated groups. The obtained intensity data were normalized by subtracting the background intensity from each oocyte size and dividing by the mean value of the control groups. All experiments were repeated at least three times and the number of analyzed oocytes are indicated in the figures.
Jo Y.J., Yoon S.B., Park B.J., Lee S.I., Kim K.J., Kim S.Y., Kim M., Lee J.K., Lee S.Y., Lee D.H., Kwon T., Son Y., Lee J.R., Kwon J, & Kim J.S. (2020). Particulate Matter Exposure During Oocyte Maturation: Cell Cycle Arrest, ROS Generation, and Early Apoptosis in Mice. Frontiers in Cell and Developmental Biology, 8, 602097.
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7

Measuring Glutathione Levels in Oocytes

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The GSH levels in oocytes were evaluated using CMF2HC (C12881, Invitrogen, Carlsbad, CA, USA). The oocytes were placed in CMF2HC, diluted with M2 medium (1:1000), and incubated at 37 °C and 5% CO2 for 30 min, protected from light, and then washed with DPBS 3 times. The samples were observed under a Nikon eclipse Ti-S microscope (ti-2U, Nikon, Tokyo, Japan).
Li Z., Zhang Y., Cao J., Xing X., Liang Y., Zhang Y., Tang X., Lin S., Wu Z., Li Z, & Huang S. (2024). Supplementation of SkQ1 Increases Mouse In Vitro Oocyte Maturation and Subsequent Embryonic Development by Reducing Oxidative Stress. Pharmaceuticals, 17(4), 455.
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8

Intracellular ROS and GSH Levels in Oocytes

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Intracellular ROS and GSH levels were detected by CM-H2DCFDA (Invitrogen, Carlsbad, CA, USA) and CMF2HC (Invitrogen), respectively. The oocytes were washed three times with DPBS-PVA and incubated in DPBS-PVA containing 5 µM CM-H2DCFDA or CMF2HC for 30 min. After incubation, oocytes were washed three times with DPBS-PVA, and fluorescence was observed under a fluorescence microscope (DMi8; Leica Microsystems, Wetzlar, Germany) with ultraviolet filters (460 nm for ROS and 370 nm for GSH). The fluorescence intensity was analyzed using ImageJ software (version 1.47; National Institutes of Health, Bethesda, MD, USA) after normalization through subtraction of the background intensity from each oocyte size. Ten to fifteen oocytes were used for each detection, and the experiment was replicated three times.
Jeong P.S., Lee S., Park S.H., Kim M.J., Kang H.G., Nanjidsuren T., Son H.C., Song B.S., Koo D.B., Sim B.W, & Kim S.U. (2020). Butylparaben Is Toxic to Porcine Oocyte Maturation and Subsequent Embryonic Development Following In Vitro Fertilization. International Journal of Molecular Sciences, 21(10), 3692.
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9

Intracellular ROS and GSH Measurement

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To measure the intracellular ROS and GSH levels, 4-cell-stage embryos were treated in 1 mL of PBS-PVA medium containing 10 mM 2′,7′-dichlorodihydrofluorescein diacetate (H2DCFDA; Invitrogen, Rochester, NY, USA) and 10 mM 4-chloromethyl-6,8-difluoro-7-hydroxycoumarin (CMF2HC; Invitrogen, Rochester, NY, USA) at 38.5 °C in an atmosphere of 5% CO2 and 100% RH for 1 h. After washing four times with PBS-PVA, the stained embryos were put into a 5 µL droplet of PBS-PVA, photographed with an inverted fluorescence microscope (Ti2eU; Nikon, Tokyo, Japan), and the fluorescence intensities of the cells were analyzed with ImageJ software (NIH).
Wang X.Q., Liu R.P., Wang J., Luo D., Li Y.H., Jiang H., Xu Y.N, & Kim N.H. (2022). Wedelolactone facilitates the early development of parthenogenetically activated porcine embryos by reducing oxidative stress and inhibiting autophagy. PeerJ, 10, e13766.
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10

OTII CD4 T Cell Conjugation Assay with B Cells and DCs

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The B:T conjugation assay (26 (link), 79 (link)) was performed as follows. Empty-GFP (CD45.11) and Mef2d-GFP (CD45.12) OTII CD4 T cells were prepared. Splenic B cells, isolated from C57BL/6J mice, were stimulated with LPS (1 μg/mL) for two days. Activated B cells were labeled with 50 μM CMF2HC (Invitrogen) for 30 minutes at room temperature, followed by a brief pulse with OVA323–339 peptide (0, 1, or 10 μg/mL) for 30 minutes at 37 °C. 1.25 × 105 empty-GFP and 1.25 × 105 Mef2d-GFP OTII CD4 T cells were co-cultured with 5.0 × 105 B cells for 20 or 30 minutes at 37 °C.
The DC:T conjugation assay was done in similar manners to the B:T conjugation assay with following modifications. Splenic DCs, after isolation with CD11c microbeads (Miltenyi Biotec) and subsequent staining with CD11c antibody for 20 minutes at 4 °C, were pulsed with OVA323–339 peptide for 2 hours at 37 °C. 2.5 × 105 DCs were co-cultured with empty-GFP and Mef2d-mAmetrine OTII CD4 T cells.
Conjugate formation of the respective OTII CD4 T cells with B cells or DCs was analyzed based on congenic molecule (CD45.11 vs. CD45.12) expression or based on GFP and mAmetrine expression among CD4 T cells in the conjugates.
Kim Y.J., Oh J., Jung S., Kim C.J., Choi J., Jeon Y.K., Kim H.J., Kim J.W., Suh C.H., Lee Y., Im S.H., Crotty S, & Choi Y.S. (2023). The transcription factor Mef2d regulates B:T synapse-dependent GC-Tfh differentiation and IL-21-mediated humoral immunity. Science immunology, 8(81), eadf2248.
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