Eclipse ti e

Manufactured by Hamamatsu Photonics

Sourced in Japan, United States

The Eclipse Ti-E is a research-grade inverted microscope platform developed by Hamamatsu Photonics. It features a modular design that enables the integration of various imaging and analysis techniques. The Eclipse Ti-E provides a stable and versatile platform for cell and tissue-based studies.

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Lab products found in correlation

Live cell imaging solution, by Thermo Fisher Scientific (1 mentions) Fluo 8h am, by AAT Bioquest (1 mentions) Orca flash 4.0 ccd camera, by Hamamatsu Photonics (1 mentions) Orca flash4.0 charge coupled device camera, by Hamamatsu Photonics (1 mentions) μ slide vi0, by Ibidi (1 mentions) Ab7753, by Abcam (1 mentions) Orca flash4.0 digital cmos camera, by Hamamatsu Photonics (1 mentions) Prolong gold antifade medium, by Thermo Fisher Scientific (1 mentions) Poly l lysine (pll), by Merck Group (1 mentions) Nis elements ar microscope imaging software, by Nikon (1 mentions) Objective heater, by Bioptechs (1 mentions) Orca flash4.0 v2 scmos, by Hamamatsu Photonics (1 mentions)

4 protocols using eclipse ti e

Fluo8H-based live-cell calcium imaging

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The Fluo8H, AM (AAT Bioquest, cat-no. 21091) was used according to the manufacturer’s instructions. In brief, Fluo8H stock solution (1.14 mol/L) was prepared by adding 1.5% Pluronic and 2.4% DMSO in PBS. Cells were incubated in EBM containing 1% FBS for 4h, Fluo8H 1:500 in starvation medium was added and cells were incubated for 30 min at 37°C, followed by 10 min at room temperature. Cells were washed twice with Live Cell Imaging Solution (Molecular Probes). Imaging of cells was performed in Live Cell Imaging Solution; compounds were added in the same volume of Live Cell Imaging Solution as the cells were cultured in, using 2x treatment solutions. Imaging was performed using a fluorescent microscope (Nikon Eclipse Ti-E, Tokyo, Japan) in combination with a Hamamatsu ORCA-Flash4.0 CCD camera (Hamamatsu, Japan). FITC fluorescence videos were acquired and quantified as stated below.

Bachmann S.B., Gsponer D., Montoya-Zegarra J.A., Schneider M., Scholkmann F., Tacconi C., Noerrelykke S.F., Proulx S.T, & Detmar M. (2019). A Distinct Role of the Autonomic Nervous System in Modulating the Function of Lymphatic Vessels under Physiological and Tumor-Draining Conditions. Cell Reports, 27(11), 3305-3314.e13.

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Dynamic Immune Cell Interactions Imaging

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imLECs were seeded into coated channeled chamber slides (μ-Slide VI^0.4; IBIDI). On the day of the assay, monolayers were preblocked with blocking antibodies for 30 min followed by the addition of 30,000 DCs, with or without blocking antibody. After 20 min, chambers were rinsed twice with media to remove nonadherent DCs. After a further 10 min equilibration at 37°C, time-lapse imaging was performed on a fluorescent microscope (Nikon Eclipse Ti-E) equipped with a Hamamatsu ORCA-Flash4.0 charge-coupled device camera (Hamamatsu) and a 20× objective (0.75 NA; Nikon). Phase contrast and FITC and APC fluorescence images were captured every 30 s for 30 min. Videos were analyzed using IMARIS software.

Arasa J., Collado-Diaz V., Kritikos I., Medina-Sanchez J.D., Friess M.C., Sigmund E.C., Schineis P., Hunter M.C., Tacconi C., Paterson N., Nagasawa T., Kiefer F., Makinen T., Detmar M., Moser M., Lämmermann T, & Halin C. (2021). Upregulation of VCAM-1 in lymphatic collectors supports dendritic cell entry and rapid migration to lymph nodes in inflammation. The Journal of Experimental Medicine, 218(7), e20201413.

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Centriole Analysis in Captured Cells

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For analysis of centrioles, the EpCAM‐captured cells were cytospun (1000g for 5 min) onto number 1.5, 13mm coverslips pretreated with poly‐L‐lysine (1mg·mL⁻¹ poly‐L‐lysine, Sigma Aldrich, St. Louis, MO, USA) for 1 h, washed 5 times with deionized water, and allowed to dry. Cells were then fixed with 100% ice‐cold methanol for 15 min. Fixed cells were blocked for 30 min in 3% BSA and 0.1% Triton X‐100 in PBS (PBSTx + BSA). Primary antibodies were incubated in PBSTx + BSA for 1 h at RT and washed three times in PBSTx, followed by secondary antibody incubation in PBSTx + BSA for 30 min at RT and two washes with PBSTx. Cells were counterstained with DAPI and mounted on glass slides with Prolong Gold antifade medium (Invitrogen). Cells were stained for centrin (Millipore, 04‐1624), pan‐cytokeratin (Abcam, ab7753), and exclusion markers (see above). Pictures were taken using the nis‐elements ar microscope imaging Software (Nikon, Melville, NY, USA) in Nikon Eclipse Ti‐E with ORCA‐Flash 4.0 Digital CMOS camera (Hamamatsu) florescent microscope with 10‐20X objectives in all the channels. Centrin foci were manually counted under 100X magnification and scored in CTCs and PBMCs.

Singh A., Denu R.A., Wolfe S.K., Sperger J.M., Schehr J., Witkowsky T., Esbona K., Chappell R.J., Weaver B.A., Burkard M.E, & Lang J.M. (2020). Centrosome amplification is a frequent event in circulating tumor cells from subjects with metastatic breast cancer. Molecular Oncology, 14(8), 1898-1909.

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Visualizing Microtubule Dynamics

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GMPCPP-stabilized microtubule seeds were prepared as described (Gell, et al., 2010 (link)) and immobilized in flow chambers using neutravidin. The final imaging buffer is BRB80 (80 mM PIPES, pH 6.8, 1 mM EGTA, 1 mM MgCl₂) supplemented with 1mM GTP, 0.1% methylcellulose 4,000 cP, 1% pluronic F-127 and 0.1 mg/ml casein. An objective heater (Bioptechs) was used to warm the chamber to 30 °C. All chambers were sealed and allowed to equilibrate on the microscope stage for 5 min prior to imaging. IRM images were acquired on a Nikon Eclipse Ti-E equipped with a Hamamatsu ORCA Flash4.0 V2 sCMOS camera every 5 s for 30 or 60 min. For depolymerization rate measurements, the frame rate used was 20 frames/s.

Chen J., Kholina E., Szyk A., Fedorov V.A., Kovalenko I., Gudimchuk N, & Roll-Mecak A. (2021). α-Tubulin Tail Modifications Regulate Microtubule Stability Through Selective Effector Recruitment, not Changes in Intrinsic Polymer Dynamics. Developmental cell, 56(14), 2016-2028.e4.

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