Upright fluorescence microscope

Manufactured by Zeiss

Sourced in Germany, United States

The Upright Fluorescence Microscope is a laboratory instrument designed for the observation and analysis of fluorescently-labeled samples. It enables the visualization and study of biological structures and processes at the microscopic level.

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Lab products found in correlation

Inverted microscope, by Zeiss (2 mentions) Metamorph, by Molecular Devices (1 mentions) Spectra id3, by Molecular Devices (1 mentions) Celltiter 96 aqueous one solution assay, by Promega (1 mentions) Alexa 488 goat anti mouse igg, by Abcam (1 mentions) Oil objective, by Zeiss (1 mentions) Triton x 100, by Merck Group (1 mentions) 4 6 diamidino 2 phenylindole (dapi), by Solarbio (1 mentions) Emccd camera, by Oxford Instruments (1 mentions) Eclipse ti e, by Nikon (1 mentions) C0065, by Solarbio (1 mentions) Dylight 488, by Thermo Fisher Scientific (1 mentions) Nbp2 13740, by Proteintech (1 mentions) Alexa fluor 594, by Thermo Fisher Scientific (1 mentions) Image pro plus software, by Media Cybernetics (1 mentions) Apoptosis assay kit, by Beyotime (1 mentions) One step tunel apoptosis assay kit, by Beyotime (1 mentions)

17 protocols using upright fluorescence microscope

Amyloid-beta Induced Neuronal Damage Assay

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A total of 4 × 10⁴ cells were cultivated on a cover glass. After 24 h, the prepared oAβ_1–42 was added and incubated for 24 h, fixed with 3% formaldehyde, punched with 0.1% Triton X-100, and placed in phosphate buffered saline with Tween (PBST) containing 5% bovine serum albumin (BSA) for blocking overnight. On the subsequent day, cells were stained with the primary antibody. After the invariable reaction overnight, cells were stained with a secondary antibody of fluorescent molecule Alexa 488 or 647. The nuclei were stained with 4′,6-diamidino-2-phenylindole (DAPI) and observed under an upright fluorescence microscope (Carl Zeiss, Jena, Germany, United Kingdom). Cell fluorescence intensity was analyzed using MetaMorph (Molecular Devices, Sunnyvale, CA, United States).

Ko C.Y., Xu J.H., Chang Y.W., Lo Y.M., Wu J.S., Huang W.C, & Shen S.C. (2022). Effects of α-Lipoic Acid on Phagocytosis of Oligomeric Beta-Amyloid1–42 in BV-2 Mouse Microglial Cells. Frontiers in Aging Neuroscience, 13, 788723.

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Bacterial Binding and Imaging with Cel-Au NCs

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After centrifuging at 8,000 rpm for 5 min, the above cultured bacterial cells were collected, washed with PBS, and incubated in the mixture of the prepared Cel-Au NCs (0.1 mL) and PBS (0.4 mL) in a shaker at 37°C for 15 min. The bacterial cultures were examined on a Zeiss upright fluorescence microscope under 605 nm.

Wang B., Fang J., Tang H., Lu S., Chen Y., Yang X, & He Y. (2023). Dual-functional cellulase-mediated gold nanoclusters for ascorbic acid detection and fluorescence bacterial imaging. Frontiers in Bioengineering and Biotechnology, 11, 1258036.

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MSC Cytocompatibility on Engineered PCL Scaffolds

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To confirm the cytocompatibility character of the etched MEW PCL mesh and engineered membranes, 1×10⁶ and 2×10⁵ MSCs were seeded per sample, respectively and cell proliferation was evaluated using MTS assay (CellTiter 96 AQueous One Solution Assay, Promega Corporation, Madison, WI, USA). At predetermined time points, the cells were incubated for 2 h and absorbance was measured in a microplate reader at 490 nm (Spectra iD3; Molecular Devices, LLC, San Jose, CA, USA). For fluorescence, the cells were fixed with 4% PFA for 30 min at RT, followed by permeabilization with 0.1% Triton X-100 in PB for 5 min at RT. The cells were then washed with PBS and blocked using 1% BSA for 30 min, followed by staining with TRITC-conjugated phalloidin and DAPI (1:1200, Millipore Sigma Company, Burlington, MA, USA) for 1 h according to the manufacturer’s instructions; samples were imaged using an upright fluorescence microscope (Carl Zeiss). SEM was done to observe cell morphology at day 5 on the membranes’ surface. The cells were fixed with 4% paraformaldehyde for 30 min, followed by washing with PBS (2×) for 5 min each. Samples were imaged using a low vacuum Tescan Rise electron microscope (Tescan USA, Inc., Warrendale, PA, USA).

Dubey N., Ferreira J.A., Daghrery A., Aytac Z., Malda J., Bhaduri S.B, & Bottino M.C. (2020). Highly Tunable Bioactive Fiber-Reinforced Hydrogel for Guided Bone Regeneration. Acta biomaterialia, 113, 164-176.

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Snail1 Expression in JNKi-Treated Cells

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JNKi treated or non-treated control 2 × 10⁴ cells were seeded onto 13 mm sterile coverslips, pre-coated with poly-L-lysine in 24 well plates, and fixed in 4% paraformaldehyde (PFA) for 15 min. Permeabilizing the cells with 0.1% Triton for 15 min at room temperature was followed by a blocking procedure for 30 min. Cells were incubated overnight at 4 °C with the primary antibody, Snail1 (20C8, Thermo Fisher, Hemel Hempstead, United Kingdom), and then with goat anti-mouse IgG (Alexa488, Abcam) secondary antibody diluted 1:500 in the washing buffer. The coverslips were then co-stained with 4’,6-diamidino-2-phenylindole (DAPI) (1.5 g/mL), and wide-field epifluorescence images were acquired at room temperature on an upright fluorescence microscope (Zeiss, Ulm, Germany). The results obtained from this experiment were from three biological repeats.

Arisan E.D., Rencuzogullari O., Keskin B., Grant G.H, & Uysal-Onganer P. (2020). Inhibition on JNK Mimics Silencing of Wnt-11 Mediated Cellular Response in Androgen-Independent Prostate Cancer Cells. Biology, 9(7), 142.

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Histological Analysis of Fractured Tibiae

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The fractured tibiae (n = 5; 3 females and 2 males) were collected for histological analyses at 7, 10, and 14 dpf. Following fixation with 10% neutral buffered formalin and decalcification using 14% ethylenediaminetetraacetic acid (EDTA), the fractured tibiae were embedded in paraffin, sectioned at 5 μm, and stained with ABH/OG. The mesenchyme, cartilage, and bone areas were measured using ImageJ software (NIH). Tissues prepared for frozen sections were fixed in 4% paraformaldehyde for 2 hours at 4°C, decalcified with 14% EDTA for 10 days, infiltrated with gradient sucrose for 3 days, embedded with Tissue-TEK OCT medium, and sectioned at a thickness of 10 μm. GFP and tdTomato fluorescence from frozen sections of fracture callus was examined by Zeiss upright fluorescence microscope.

Xiao D., Fang L., Liu Z., He Y., Ying J., Qin H., Lu A., Shi M., Li T., Zhang B., Guan J., Wang C., Abu-Amer Y, & Shen J. (2024). DNA methylation–mediated Rbpjk suppression protects against fracture nonunion caused by systemic inflammation. The Journal of Clinical Investigation, 134(3), e168558.

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Lung Epithelial Cell Ablation Dynamics

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Lungs from control and Yap^f/f; Shh^Cre/+; ROSA26^mTmG/+ embryos at 12.5 dpc were collected in cold PBS and mounted onto concavity slides with coverslips. Epithelial cells were imaged through their expression of GFP (from the ROSA26^mTmG/+ allele) with a 40× oil objective (NA = 1.2, Zeiss). A 10 μm span of epithelial cells in the left lobe was targeted for 2 s by a Chameleon 915 nm two-photon laser (~700 mW) mounted on a custom-built Zeiss upright fluorescence microscope. Immediately following ablation, time-series images of epithelial cells were taken at the frame rate of 1 s for up to 50 s. The movements of the epithelial cells from 0 to 5 s were tracked using the manual tracking plug-in of ImageJ (NIH).

Lin C., Yao E., Zhang K., Jiang X., Croll S., Thompson-Peer K, & Chuang P.T. (2017). YAP is essential for mechanical force production and epithelial cell proliferation during lung branching morphogenesis. eLife, 6, e21130.

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Immunofluorescence Analysis of NAT10 and RNPS1

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The cell climbing sheets were fixed with 4% paraformaldehyde in PBS overnight and stained with the following primary antibody: anti-NAT10 (Santa Cruz, sc-271770 1:100) & anti-RNPS1 (Proteintech, 10555-1-AP, 1:100) after permeated with 1% Triton™ X-100 (Sigma-Aldrich, X-100). Then, the antigens were visualized by the corresponding secondary antibody conjugated with DyLight 488 and Fluor 594. The nuclei were counterstained with DAPI (Solarbio, C0065) at 1:1 000 for 1 min. Images were captured with an upright fluorescence microscope (ZEISS, Germany).

Wang X., Ling R., Peng Y., Qiu W, & Chen D. (2024). RNPS1 stabilizes NAT10 protein to facilitate translation in cancer via tRNA ac4C modification. International Journal of Oral Science, 16, 6.

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Identifying ILC2s in Nasal Mucosa

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For immunofluorescence double staining, the wrapped nasal mucosa was fixed in 4% neutral buffered formalin and decalcified in an EDTA solution. Slices of 4 μm thickness were obtained and dewaxed in xylene, followed by ethanol gradient dehydration and a PBS wash. Thy-1 (CD90) and ST2 antibodies (Abcam, Cambridge, UK) were used to determine the number of ILC2s. Photo capture was performed using an upright fluorescence microscope (Zeiss, Germany).

Xie Y., Zhang Y., Zhu T., Ma J., Duan C., Yang L., Wang T., Zhuang R., Bian K, & Lu L. (2022). CD226 Deficiency Alleviates Murine Allergic Rhinitis by Suppressing Group 2 Innate Lymphoid Cell Responses. Mediators of Inflammation, 2022, 1756395.

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Quantifying ILC2s in Nasal Mucosa

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Five mice from each group were randomly selected for immunofluorescence double staining to detect the number of ILC2s. The nasal mucosa wrapped by nasal bone was fixed in 4% neutral buffered formalin, then decalcified in EDTA solution. Three-micrometer slices were obtained and dewaxed in xylene, followed by ethanol gradient dehydration and washed in phosphate buffer solution (PBS). CD90 (Thy1) and ST2 antibodies (Abcam, Cambridge, UK) were employed to stain the aimed protein according to the manufacturer protocol. Photo capture was performed using upright fluorescence microscope (Zeiss, Germany).

Hu B., Wang Y., Zheng G., Zhang H, & Ni L. (2021). Effect of parasympathetic inhibition on expression of ILC2 cells in a mouse model of allergic rhinitis. The World Allergy Organization Journal, 14(9), 100582.

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Quantitative microscopy of bacterial cells

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For dose-response experiments, static images were acquired by imaging cells on a 1% agarose pad made using the medium cells had been grown in using an upright fluorescence microscope (Zeiss, Thornwood, NY, USA) equipped with a Plan-Apo 1.4 100x phase objective, conventional epifluorescence filter set, and 1024 × 1024 back illuminated EM-CCD camera (Andor, South Windsor, CT, USA), producing an effective pixel size of 66 nm. Dynamic experiments were imaged using a Nikon Eclipse TiE enclosed in a temperature controlled chamber (Haison Technology, Taipei, Taiwan) set to 30°C in the case of C. crescentus and 37°C in the case of E. coli using a Plan-Apo 1.45 100x phase objective and 1024 × 1024 back illuminated EM-CCD camera (Andor, South Windsor, CT, USA), producing an effective pixel size of 86 nm. E. coli nutritional upshift experiments were performed using an ONIX Microfluidic Perfusion Platform (CellASIC, Hayward, CA, USA). C. crescentus perfusion experiments were performed using a low profile RC-31 flow chamber (Warner, Hamden, CT, USA). See Supplemental Experimental Procedures for further details.

Harris L.K, & Theriot J.A. (2016). Relative Rates of Surface and Volume Synthesis Set Bacterial Cell Size. Cell, 165(6), 1479-1492.

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