Dmi 400b fluorescence microscope

Manufactured by Leica

The DMI 400B is a fluorescence microscope designed for life science research applications. It is capable of visualizing and analyzing fluorescently-labeled samples. The DMI 400B offers high-quality optics and illumination systems to provide clear and detailed images of fluorescent specimens.

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Lab products found in correlation

Alexa fluor 488 donkey anti rabbit igg secondary antibody, by Thermo Fisher Scientific (1 mentions) Anti mdm2, by Thermo Fisher Scientific (1 mentions) Alexa fluor 568 donkey anti mouse igg secondary antibody, by Thermo Fisher Scientific (1 mentions) Anti e cadherin, by Santa Cruz Biotechnology (1 mentions) Anti p smad3, by Cell Signaling Technology (1 mentions) Anti mdm2, by Santa Cruz Biotechnology (1 mentions) Bovine serum albumin (bsa), by Merck Group (1 mentions) A21206, by Thermo Fisher Scientific (1 mentions) Fitc conjugated secondary antibody, by Thermo Fisher Scientific (1 mentions) Nunc lab tek, by Thermo Fisher Scientific (1 mentions) Anti yap antibody, by Cell Signaling Technology (1 mentions) Sc 74495, by Santa Cruz Biotechnology (1 mentions)

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Fluorescence microscope (2465 citations)

5 protocols using dmi 400b fluorescence microscope

Immunofluorescent detection of MDM2 and pSmad3

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Cells transfected with pCMVβ-myc3-MDM2 plasmid or empty vector were plated in 6-well plates. Then cells were fixed with 4% paraformaldehyde in PBS for 15 min, and blocked with 3% bovine serum albumin (Sigma-Aldrich, A7906) and 0.3% Triton X-100 in PBS for 1 h at room temperature. MDM2 and p-Smad3 was probed with primary antibodies anti-MDM2 (1 : 50) and anti-pSmad3 (1 : 150) in PBS overnight at 4 °C at room temperature and Alexa Fluor 488 donkey anti- rabbit IgG secondary antibody (Invitrogen, A-11008) or Alexa Fluor 568 donkey anti-mouse IgG secondary antibody (Invitrogen, A-10037) diluted in 1 : 500 in PBS for 1 h at 37 °C. Nuclei were visualised by staining with DAPI (4′ 6-diamidino-2-phenylindole; Sigma-Aldrich, D9542). Cells were than washed thrice with PBS and imaged with Leica DMI 400B fluorescence microscope.
anti-MDM2 (Cat# sc-965), anti-E-cadherin (Cat# sc-7870) were purchased from Santa Cruz (Santa Cruz), and anti-p-Smad3 (Cat# 9520) was from Cell Signaling Technology (Beverly, MA, USA).

Chen Y., Wang D.D., Wu Y.P., Su D., Zhou T.Y., Gai R.H., Fu Y.Y., Zheng L., He Q.J., Zhu H, & Yang B. (2017). MDM2 promotes epithelial–mesenchymal transition and metastasis of ovarian cancer SKOV3 cells. British Journal of Cancer, 117(8), 1192-1201.

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Quantifying DNA Damage in HepG2 Cells

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HepG2 cells were exposed to Q6 (50 μM) or TPZ (100 μM) for 1 h. For γ-H2AX foci assessment, the cells were fixed in 4% paraformaldehyde in PBS for 20 min at room temperature followed by three rinses in PBS and permeabilization in 0.1% Triton-X100 (in PBS) for 10 min at 4°C. Next, cells were blocked by 4% BSA and incubated with primary rabbit monoclonal antibody against γH2AX (1:200) for 10 h at 4°C. Following two rinses with PBS, cells were incubated for 1 hour with FITC-conjugated secondary antibody (1:200, A-21206, Invitrogen), and subjected to with DAPI staining (1 μg/mL) and imaged with Leica DMI 400B fluorescence microscope.

Chang L., Liu X., Wang D., Ma J., Zhou T., Chen Y., Sheng R., Hu Y., Du Y., He Q., Yang B, & Zhu H. (2015). Hypoxia-Targeted Drug Q6 Induces G2-M Arrest and Apoptosis via Poisoning Topoisomerase II under Hypoxia. PLoS ONE, 10(12), e0144506.

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Cell Proliferation Assay Using BrdU

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Cells were seeded in 6-well plates at a cell density of 1 × 10⁴ cells/well in reduced serum and growth factor-containing DMEM and treated with the reagents as described above. After treatment for 24 h, BrdU was added at a final concentration of 30 µg/L in the medium, and incubated with the cells for 4 h. The cells were fixed with methanol containing 0.04 % H₂O₂ for 15 min; they were then incubated in 2 N hydrochloric acid at 37 °C for 1 h. Subsequently, the cells were blocked in 5 % bovine serum albumin (BSA) for 20 min at room temperature, and were then incubated overnight with the BrdU antibody (1:200 dilution) at 4 °C. After the first antibody was washed off, the cells were incubated with goat anti-mouse IgG-FITC (1:50 dilution) for 30 min at room temperature lucifugally. This was followed by excitation at 494 nm, after which BrdU-positive cells were counted under the Leica DMI 400B fluorescence microscope under 200× magnification.

Wang Y., Yin D., Xu C., Wang K., Zheng L, & Zhang Y. (2016). Roxarsone induces angiogenesis via PI3K/Akt signaling. Cell & Bioscience, 6, 54.

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Evaluating Tumor Hypoxia and YAP Localization

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HepG2 cells were collected and inoculating subcutaneously into nude mice. After the tumor size reached a mean group size of 300–500 mm³, the hypoxia probe Hypoxyprobe^™-1 (Pimonidazole Hydrochloride, PIMO) (60 mg/kg) was injected intraperitoneally to indicate tumor hypoxic regions. One hour later, the tumors were removed and cut into frozen slices. For detection of the intratumor and subcellular localization of YAP, the cells grown in glass bottom dishes (Thermo Scientific Nunc Lab-Tek) as well as the frozen slices were fixed with 4% paraformaldehyde, and blocked with 5% BSA, and incubated with anti-YAP antibody (Cell Signaling Technology, Beverly, Mass) and FITC-conjugated secondary antibody (Invitrogen). The slides were than stained with DAPI and imaged with Leica DMI 400B fluorescence microscope.

Dai X.Y., Zhuang L.H., Wang D.D., Zhou T.Y., Chang L.L., Gai R.H., Zhu D.F., Yang B., Zhu H, & He Q.J. (2016). Nuclear translocation and activation of YAP by hypoxia contributes to the chemoresistance of SN38 in hepatocellular carcinoma cells. Oncotarget, 7(6), 6933-6947.

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Osteocalcin Immunostaining in Cells

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Cells grown and treated in 96-well plates were fixed with 4% formaldehyde in PBS for 15 min, washed two times with PBS, and blocked with 5% FBS and 1% Triton X-100 in PBS for 1 h at room temperature. Osteocalcin were detected with antibody (SC-74495) from Santa Cruz Biotechnology Inc. and PE-conjugated secondary antibody diluted 1∶100 in PBS for 2 h at room temperature. Cells were then washed three times in PBS and imaged with a Leica DMI 400B fluorescence microscope.

Zhang N., Ying M.D., Wu Y.P., Zhou Z.H., Ye Z.M., Li H, & Lin D.S. (2014). Hyperoside, a Flavonoid Compound, Inhibits Proliferation and Stimulates Osteogenic Differentiation of Human Osteosarcoma Cells. PLoS ONE, 9(7), e98973.

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