Image it live plasma membrane and nuclear labeling kit

Manufactured by Thermo Fisher Scientific

Sourced in United States

The Image-iT LIVE plasma membrane and nuclear labeling kit is a fluorescence-based product that enables simultaneous visualization of the plasma membrane and nuclei in live cells. The kit provides a set of optimized reagents for staining the plasma membrane and nucleus, facilitating the observation of these cellular structures under live-cell imaging conditions.

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Lab products found in correlation

1 phenyl 2 thiourea ptu, by Merck Group (2 mentions) Tricaine, by Merck Group (2 mentions) Hanks balanced salt solution (hbss), by Thermo Fisher Scientific (2 mentions) Formaldehyde, by Merck Group (2 mentions) Stellaris 5 confocal microscope, by Leica (1 mentions) Poly l lysine (pll), by Solarbio (1 mentions) Cell imaging coverglass, by Eppendorf (1 mentions) Lsm 780, by Zeiss (1 mentions) Vectashield antifade mounting medium, by Vector Laboratories (1 mentions) Lipofectamine 2000 transfection reagent, by Thermo Fisher Scientific (1 mentions) Alexa fluor 488, by Thermo Fisher Scientific (1 mentions) Ab94396, by Abcam (1 mentions) Fluorescence microscope, by Leica (1 mentions) Hanks balanced salt solution (hbss), by Corning (1 mentions) Hanks balanced salt solution (hbss), by Leica (1 mentions) Glass coverslip, by Thermo Fisher Scientific (1 mentions) Hanks balanced salt solution (hbss), by Merck Group (1 mentions) Hanks balanced salt solution (hbss), by Nikon (1 mentions)

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Image-iT (18 citations)

7 protocols using image it live plasma membrane and nuclear labeling kit

Zebrafish Tumor Cell Tracking

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Zebrafish (AB, wild type [WT], 48-h embryo) embryos were obtained from the Core Facilities, Zhejiang University, School of Medicine, cultured in Petri dishes with an appropriate amount of water and placed in a 28 °C incubator under constant temperature and light. Then, 0.003% 1-phenyl-2-thiourea (PTU) (Sigma) was added within 24 h after the embryo was produced. In total, 200 SiHa cells were harvested after transfection and resuspended in PBS. Plasma membranes were stained with red-fluorescent Alexa Fluor® 594 wheat germ agglutinin of the Image-iT™ LIVE Plasma Membrane and Nuclear Labeling Kit (Molecular Probes, USA, I34406). Zebrafish embryos were anesthetized with 0.016% tricaine (Sigma). Then, the cells were injected into the yolk sac of zebrafish embryos. Then, we obtained images with a fluorescence microscope at 48 h after injection. The area of red fluorescence in the zebrafish yolk sac was calculated using ImageJ software [33 ]. Next, the zebrafish embryos were anesthetized with an overdose of tricaine.

Zhang Y., Zhao L., Yang S., Cen Y., Zhu T., Wang L., Xia L., Liu Y., Zou J., Xu J., Li Y., Cheng X., Lu W., Wang X, & Xie X. (2020). CircCDKN2B-AS1 interacts with IMP3 to stabilize hexokinase 2 mRNA and facilitate cervical squamous cell carcinoma aerobic glycolysis progression. Journal of Experimental & Clinical Cancer Research : CR, 39, 281.

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Zebrafish Membrane Labeling and Cell Injection

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Zebrafish (AB, wild type [WT], 48-h embryo) embryos were obtained from the Core Facilities, Zhejiang University, School of Medicine, cultured in Petri dishes with an appropriate amount of water and placed in a 28°C incubator under constant temperature and light. Then, 0.003% 1-phenyl-2-thiourea (PTU) (Sigma) was added within 24 h after the embryo was produced. In total, 200 SiHa cells were harvested after transfection and resuspended in PBS. Plasma membranes were stained with red-fluorescent Alexa Fluor® 594 wheat germ agglutinin of the Image-iT™ LIVE Plasma Membrane and Nuclear Labeling Kit (Molecular Probes, USA, I34406). Zebrafish embryos were anesthetized with 0.016% tricaine (Sigma). Then, the cells were injected into the yolk sac of zebrafish embryos. Then, we obtained images with a fluorescence microscope at 48 h after injection. The area of red fluorescence in the zebrafish yolk sac was calculated using ImageJ software [33] . Next, the zebrafish embryos were anesthetized with an overdose of tricaine.

Zhang Y., Zhao L., Yang S., Cen Y., Zhu T., Wang L., Xia L., Liu Y., Zou J., Xu J., Li Y., Cheng X., Lu W., Wang X., & Xie X. (2020). CircCDKN2B-AS1 interacts with IMP3 to stabilize Hexokinase 2 mRNA and facilitate cervical squamous cellcarcinoma aerobic glycolysis progression.

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Imaging of Synchronized Plasmodium Parasites

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Live-cell imaging was performed as described (85 (link)). Briefly, after synchronization with 5% sorbitol at the early ring stage (3 hpi), parasites were allowed to mature; schizonts (42 hpi) were purified from uninfected RBCs using 40%/70% Percoll as described above and treated with 10 μM E64 cysteine protease inhibitor (ThermoFisher) for 3 to 4 h to prevent egress; and the parasites were incubated with fresh RBCs at 2% hematocrit in RPMI complete medium containing dyes from the Image-IT live plasma membrane and nuclear labeling kit (Thermofisher) at 37°C for 10 min. The labeled parasites were washed twice in PBS and then plated on a cell imaging cover glass (Eppendorf, Hamburg, Germany) coated with poly-l-lysine (Solarbio, Beijing, China). Live-cell imaging of the parasites was performed on a Leica Stellaris 5 confocal microscope. The sample chamber was maintained at 37°C and supplied with a humidified atmosphere under 5% CO₂.

He L., Qiu Y., Pang G., Li S., Wang J., Feng Y., Chen L., Zhu L., Liu Y., Cui L., Cao Y, & Zhu X. (2023). Plasmodium falciparum GAP40 Plays an Essential Role in Merozoite Invasion and Gametocytogenesis. Microbiology Spectrum, 11(3), e01434-23.

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Subcellular Localization of SOX5 in HEK-293 Cells

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HEK-293 cells (ATCC® CRL-1573™) were plated on glass coverslips and transfected with pKTol2C-SOX5 plasmids (2 μg) and Lipofectamine 2000 Transfection Reagent (Thermo Fisher Scientific). Two days later, they were stained using Image-IT™ LIVE Plasma Membrane and Nuclear Labeling Kit (Thermo Fisher, I34406), fixed in 4% paraformaldehyde, permeabilized with 0.1% Triton X-100 in PBS (PBST), and blocked in PBST supplemented with 1% BSA and 22.5 mg/ml glycine. They were then incubated with rabbit polyclonal SOX5 antibody (1:200, Abcam, ab94396), followed by goat anti-rabbit antibody (1:500, Alexa Fluor 488, Invitrogen, A27034). After placing DAPI-containing Vectashield antifade mounting medium (Vector Laboratories), cells were imaged by confocal laser scanning microscopy (Zeiss LSM 780; 100× objective).

Zawerton A., Mignot C., Sigafoos A., Blackburn P.R., Haseeb A., McWalter K., Ichikawa S., Nava C., Keren B., Charles P., Marey I., Tabet A.C., Levy J., Perrin L., Hartmann A., Lesca G., Schluth-Bolard C., Monin P., Dupuis-Girod S., Guillen Sacoto M.J., Schnur R.E., Zhu Z., Poisson A., Chehadeh S.E., Alembik Y., Bruel A.L., Lehalle D., Nambot S., Moutton S., Odent S., Jaillard S., Dubourg C., Hilhorst-Hofstee Y., Barbaro-Dieber T., Ortega L., Bhoj E.J., Masser-Frye D., Bird L.M., Lindstrom K., Ramsey K.M., Narayanan V., Fassi E., Willing M., Cole T., Salter C.G., Akilapa R., Vandersteen A., Canham N., Rump P., Gerkes E.H., Wassink-Ruiter K.J., Bijlsma E., Hoffer M.J., Vargas M., Wojcik A., Cherik F., Francannet C., Rosenfeld J.A., Machol K., Scott D.A., Bacino C.A., Wang X., Clark G.D., Bertoli M., Zwolinski S., Thomas R.H., Akay E., Chang R.C., Bressi R., Russo R.S., Srour M., Russell L., Goyette A.M., Dupuis L., Mendoza-Londono R., Karimov C., Joseph M., Nizon M., Cogné B., Kuechler A., Piton A., Klee E.W., Lefebvre V., Clark K.J, & Depienne C. (2019). Widening of the genetic and clinical spectrum of Lamb–Shaffer syndrome, a neurodevelopmental disorder due to SOX5 haploinsufficiency. Genetics in medicine : official journal of the American College of Medical Genetics, 22(3), 524-537.

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Visualizing SWCNT Uptake in BEAS-2B Cells

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BEAS-2B cells were seeded overnight on glass coverslips in a 12 well plate at 1.5 × 10⁵ cell/well and exposed to 50 μg/ml pristine or treated SWCNTs for 24 h. Subsequently, the exposed cells were washed twice with Hank’s balanced salt solution (HBSS, Corning), and fixed with 4% formaldehyde (Sigma-Aldrich) for 15 min at 37 °C. Upon fixation, the exposed cells were washed again 3 times with HBSS and stained with 3 μg/ml Alexa Fluor 594 wheat germ agglutinin (WGA) and 2 μM Hoechst 33,342 (Image-iT LIVE plasma membrane and nuclear labeling kit, Life Technologies) respectively for 10 min. Lastly, the coverslips with stained cells were washed twice with HBSS, mounted on glass slides, and imaged using a fluorescence microscope (Leica) and a 40 × objective.

Eldawud R., Wagner A., Dong C., Stueckle T.A., Rojanasakul Y, & Dinu C.Z. (2017). Carbon nanotubes physicochemical properties influence the overall cellular behavior and fate. NanoImpact, 9, 72-84.

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PLACC900 Exposure on BEAS-2B Cells

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BEAS-2B cells were seeded on glass coverslips (15 mm diameter; Fisher Scientific) in a 12 well plate at a density of 1.5 × 10⁵ cells/ml overnight. The cells were subsequently exposed to 100, 300, or 500 µg/ml PLACC900 dispersed in media as previously described. After 24 h, the media was removed and the cells were washed two times with Hank’s Balanced Salt Solution (HBSS) (Corning, Inc.), fixed with 4% formaldehyde (Sigma-Aldrich) for 15 min and at 37 °C, and subsequently washed 3 more times with HBSS to remove any remaining formaldehyde. The cells’ plasma membranes and nuclei were then stained with 3 µg/ml Alexa Fluor 594 wheat germ agglutinin (WGA) and 2 µM Hoechst 33342 (Image-iT LIVE Plasma Membrane and Nuclear Labeling Kit, Life Technologies), respectively, both dispersed in HBSS, for 10 min and at 4 °C. After incubation, cells were washed 2 times with HBSS, the cover slides were mounted on glass coverslips, and imaged under a Nikon Inverted Microscope Eclipse Ti Series (Nikon) and a 40X objective. The NIS-Elements BR 3.1 software was used to analyze the size and morphology of the cells.

Wagner A., White A.P., Tang M.C., Agarwal S., Stueckle T.A., Rojanasakul Y., Gupta R.K, & Dinu C.Z. (2018). Incineration of Nanoclay Composites Leads to Byproducts with Reduced Cellular Reactivity. Scientific Reports, 8, 10709.

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Cellular Localization of AGuIX Nanoparticles

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The experiment was performed using a LEICA confocal microscope system. The cells were thermostatically controlled and regulated in CO₂. B16F10 cells were grown in 4-well LabTek chambers I at a density of 10,000 cells/well. For localization studies, cells were incubated with 0.6 mg/L with AGuIX® functionalized with FITC as a fluorescent marker for 1 hour. After incubation, cells were rinsed and then incubated with 5 µg/ml fluorescent Alexa Fluor 594 wheat germ agglutinin (Image-iT LIVE Plasma Membrane and Nuclear labeling kit, Life Technologies) for highly selective staining of the plasma membrane for 10 minutes at 37°C and 5 % CO₂. Afterwards, cells were suspended in Hank's balanced salt solution (HBSS) for imaging. FITC was excited at 499 nm, and the fluorescence emission was detected at 520 nm. Alexa Fluor 594 was excited at 592 nm, and emission was detected at 620 nm.

Kotb S., Detappe A., Lux F., Appaix F., Barbier E.L., Tran V.L., Plissonneau M., Gehan H., Lefranc F., Rodriguez-Lafrasse C., Verry C., Berbeco R., Tillement O, & Sancey L. (2016). Gadolinium-Based Nanoparticles and Radiation Therapy for Multiple Brain Melanoma Metastases: Proof of Concept before Phase I Trial. Theranostics, 6(3), 418-427.

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