Epifluorescent microscope

Manufactured by Nikon

Sourced in Japan

The Epifluorescent microscope is a type of optical microscope that uses fluorescence to produce images of samples. It illuminates the specimen with light of a specific wavelength, causing fluorescent molecules within the sample to emit light at a different wavelength, which is then detected and used to create an image.

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Lab products found in correlation

Openlab software, by PerkinElmer (2 mentions) Tff 3, by Santa Cruz Biotechnology (1 mentions) Anti ng2, by Merck Group (1 mentions) Anti cd31, by BD (1 mentions) Anti α sma, by Merck Group (1 mentions) Isolectin b4, by Vector Laboratories (1 mentions) Qimaging camera, by Nikon (1 mentions) Scmos camera, by Oxford Instruments (1 mentions) 4 6 diamidino 2 phenylindole (dapi), by Biotium (1 mentions) Proteostat aggresome dye, by Enzo Life Sciences (1 mentions) β galactosidase staining kit, by Cell Signaling Technology (1 mentions) Poly d lysine, by Merck Group (1 mentions) Laminin, by Merck Group (1 mentions) Dsu confocal microscope, by Olympus (1 mentions) Glial fibrillary acidic protein (gfap), by Merck Group (1 mentions) A1 confocal microscope, by Nikon (1 mentions)

Spelling variants of this product

Eclipse TI epifluorescent microscope (6 citations) Inverted epifluorescent microscope (4 citations) Ni-E epifluorescent microscope (3 citations) Widefield epifluorescent microscope (3 citations) Eclipse E800 epifluorescent microscope (2 citations) Ti-2e Epifluorescent microscope (2 citations) 90i epifluorescent microscope (2 citations) Nikon epifluorescent microscope 80i (2 citations) TE2000 epifluorescent microscope (2 citations) TE2000-U inverted epifluorescent microscope (2 citations)

17 protocols using epifluorescent microscope

Colonic Tissue Immunostaining and FISH

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Formalin-fixed paraffin-embedded colonic tissues were sectioned (6 μm) and analyzed via immunofluorescence and FISH. For FISH analysis, tissue sections were hybridized with 10 ng/mL of a universal bacterial 16S fluorescent rRNA probe, according to previously published protocol [99 (link)]. For immunofluorescence, tissue sections were incubated with anti-CR LPS (Biotech Laboratories, Ottawa, Canada), biotinylated goat anti-GFP (GeneTex, Irvine, USA), anti-MBD-3 (Santa Cruz Biotechnology, Dallas, USA), anti-TFF3 (Santa Cruz Biotechnology, Dallas, USA) overnight at 4°C.
Confluent Caco-2 monolayers were fixed with 100% methanol and permeabilized with 0.1% Triton 1% BSA solution in PBS. Primary antibodies were used to detect β defensin-2 (Santa Cruz Biotechnology, Dallas, USA) and TFF3 (Santa Cruz Biotechnology, Dallas, USA). The selectivity of these antibodies has been previously validated [61 (link), 100 (link)–102 (link)]. Caco2 cells are known to produce low levels of TFF3, and the use of Caco2 enterocytes to detect modulation of TFF3 release has been previously validated [103 (link)–105 (link)]. Images were acquired using a Nikon epifluorescent microscope, and ImageJ was used for microscopic image analysis.

Manko A., Motta J.P., Cotton J.A., Feener T., Oyeyemi A., Vallance B.A., Wallace J.L, & Buret A.G. (2017). Giardia co-infection promotes the secretion of antimicrobial peptides beta-defensin 2 and trefoil factor 3 and attenuates attaching and effacing bacteria-induced intestinal disease. PLoS ONE, 12(6), e0178647.

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Quantitative Analysis of Pericyte Coverage

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Explant tissues were cultured, washed, and stained using standard protocols as mentioned above. For the pericyte coverage analysis, the endothelium in the metatarsal assay was stained with an anti-CD31 (BD Pharmingen, 553370) antibody and in the aortic ring assay with Isolectin B4 (Vector Labs, FL-1201). Pericytes were stained with anti-αSMA (Sigma, C6198) and anti-NG2 (Millipore, AB5320) antibodies. Single images were taken using a 20 × objective focussed on the plane of vessels using a Nikon epifluorescent microscope. For quantitative analysis, using NIS Elements Software, single thresholds were calculated for each individual channel (endothelium, pericytes) and the intersection of the endothelium covered by pericytes. The analysis was blinded to treatments and was performed on masked images with different thresholds used per image as required to avoid saturation. Pericyte coverage was then calculated as % (Intersection/Endothelium), and the values were normalised to the average of the internal control (vehicle/PBS) for every experiment. The normalised values of at least three independent experiments were pooled for statistical analysis. Outliers were eliminated using the average ± 2SD mean.

Dritsoula A., Dowsett L., Pilotti C., O’Connor M.N., Moss S.E, & Greenwood J. (2022). Angiopathic activity of LRG1 is induced by the IL-6/STAT3 pathway. Scientific Reports, 12, 4867.

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Quantifying Anaerobic Microbiome in Sewage Sludge

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The molecular analysis aimed at determining the percentage content of anaerobic microorganisms in sewage sludge after AD by using the fluorescent in situ hybridization (FISH) technique. Four molecular probes were used for hybridization: a universal probe for Bacteria EUB338, a universal probe for Archaea ARC915, a probe oriented for Methanosarcinaceae MSMX860, and a probe oriented for Methanosaeta MX825. The samples were analyzed under an epifluorescent microscope (Nikon, Tokyo, Japan) with a 100× lens and total magnification at 1000×. The population numbers of the studied microorganisms were counted from cells stained with DAPI using Image Processing and Analysis in Java (ImageJ – https://imagej.net/software/imagej/) (LOCI, University of Wisconsin, Madison, WI, USA) [73 ].

Kazimierowicz J., Dębowski M, & Zieliński M. (2023). Biohythane Production in Hydrogen-Oriented Dark Fermentation of Aerobic Granular Sludge (AGS) Pretreated with Solidified Carbon Dioxide (SCO2). International Journal of Molecular Sciences, 24(5), 4442.

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Quantifying Synapse Formation in C. elegans

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Analysis was carried out on live animals at 40× magnification using a Nikon epifluorescent microscope and a Q-imaging camera. Animals were anesthetized using 1% (v/v) 1-phenoxy-2-propanol in M9 buffer. Synapse formation defects were quantified by collecting images of juIs1 (P_unc-25SNB-1::GFP) and manually scoring puncta numbers in Adobe Photoshop. Dorsal cord lengths were determined in µmeters using Q-imaging software. For each genotype 20 or more worms were analyzed from at least 3 independent experiments. Both bar-1 and pop-1 mutants displayed stereotyped, reproducible gaps in their dorsal cords (data not shown). Care was taken to avoid collecting images at these locations. Axon termination defects were visualized using muIs32 (P_mec-7GFP) and manually scored. For all genetic analysis on axon termination, averages are shown for data collected from 5–8 independent counts of 20–30 PLM neurons from young adult worms. For transgenic analysis, data shown is an average of 4 or more transgenic lines for each genotype.

Tulgren E.D., Turgeon S.M., Opperman K.J, & Grill B. (2014). The Nesprin Family Member ANC-1 Regulates Synapse Formation and Axon Termination by Functioning in a Pathway with RPM-1 and β-Catenin. PLoS Genetics, 10(7), e1004481.

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Live-Cell Imaging of Inducible Inclusion Formation

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Infected monolayers of Cos7 cells grown in a glass bottom 24 well plate were induced at 16 hpi with either 0.5mM theophylline only or the indicated concentrations of theophylline and anhydroTetracycline. Plates were imaged immediately upon induction.
Live cell imaging was achieved using an automated Nikon epifluorescent microscope equipped with an Okolab (http://www.oko-lab.com/live-cell-imaging) temperature controlled stage and an Andor Zyla sCMOS camera (http://www.andor.com). Images were taken every fifteen minutes for a further 36 hours. Multiple fields of view of multiple wells were imaged. The fluorescence intensity of each inclusion over time was tracked using the ImageJ plugin Trakmate [18 (link)]. and the results were averaged and plotted using python and matplotlib [19 (link)].

Grieshaber N.A., Chiarelli T.J., Appa C.R., Neiswanger G., Peretti K, & Grieshaber S.S. (2022). Translational gene expression control in Chlamydia trachomatis. PLoS ONE, 17(1), e0257259.

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Detecting Protein Aggregation in Cells

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ProteoStat Aggresome dye (Enzo Life Sciences, PA) was used to detect misfolded and aggregated proteins in cells (Shen et al., 2011 (link)). Cells were grown on multi-chamber glass bottom slides and treated with vehicle or patulin. After 24 h, cells were fixed in 4% paraformaldehyde for 30 min at room temperature and permeabilized with 0.5% Triton X-100 in 1 × assay buffer for 30 min on ice. Cells were washed with 1 × assay buffer for two times and stained for 30 min at room temperature with ProteoStat Aggresome dye. Nuclei were stained with 4′,6-diamidino-2-phenylindole (DAPI; Biotium) for 5 min. Stained cells were examined with a Nikon epifluorescent microscope using a Texas Red filter set for the ProteoStat dye, and UV for blue fluorescence for DAPI, respectively. Images were acquired using a 63 × objective lens with a Spot RT3 digital camera and Adobe Photoshop CS was used to layer the captured images. Fluorescence intensity per cell was quantified using NIH ImageJ analysis software to obtain mean corrected total cell fluorescence (CTCF) of 8–10 readings/area per high power field, and CTCF was calculated as: Integrated density– (Area of selected cell x mean background fluorescence).

Han J.J., Nguyen C.D., Thrasher J.P., DeGuzman A, & Chan J.Y. (2022). The Nrf1 transcription factor is induced by patulin and protects against patulin cytotoxicity. Toxicology, 471, 153173.

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Quantifying Cellular Changes in Brain Injury

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Following double immunofluorescence staining, numbers of positively stained cells in the intramatrix zone (IMZ) and lesion boundary zone (LBZ) on day 7 (D7), D14, D21, and D28 were counted manually in three to five different fields per section of each rat brain (using an eyepiece grid covering an area of 0.0625 mm²) by an individual who was blinded to the experimental design. Vessels and blood cells were excluded. Sections were observed, and images were acquired using a Nikon epifluorescent microscope. Total cells in the IMZ and LBZ were visualized by DAPI staining at 20x magnification using Openlab software (Improvision, Cambridge, MA). Ki67⁺ cells were visualized as fluorescent red (DL 549), and NeuN⁺ cells were visualized as fluorescent green (Alexa 488) in the IMZ and LBZ at 20x, while double-positive cells were visualized as yellow. For double immunofluorescence staining, double-positive cells were also manually counted in the same manner. Finally, we present the results as the number of immunopositive cells per field.

Huang K.F., Hsu W.C., Hsiao J.K., Chen G.S, & Wang J.Y. (2014). Collagen-Glycosaminoglycan Matrix Implantation Promotes Angiogenesis following Surgical Brain Trauma. BioMed Research International, 2014, 672409.

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Assessing Cisplatin and M6620 Impact

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Cells were seeded in six-well plates on the day prior to treatment. Cells were subsequently left untreated, treated with 500 nmol/l cisplatin for 2 h, 500 nmol/l M6620 for 4 h or 500 nmol/l cisplatin + 500 nmol/l M6620. Cells were allowed to grow for 6 days after which cells were fixed and subsequently incubated with X-gal substrate overnight at 37°C as per the manufacturer's instructions using a β-galactosidase staining kit (Cell Signaling Technologies). Experiments were performed twice, and images taken on a Nikon epifluorescent microscope using a 20× air objective. Images were equally adjusted for presentation purposes.

Heyza J.R., Ekinci E., Lindquist J., Lei W., Yunker C., Vinothkumar V., Rowbotham R., Polin L., Snider N.G., Van Buren E., Watza D., Back J.B., Chen W., Mamdani H., Schwartz A.G., Turchi J.J., Bepler G, & Patrick S.M. (2023). ATR inhibition overcomes platinum tolerance associated with ERCC1- and p53-deficiency by inducing replication catastrophe. NAR Cancer, 5(1), zcac045.

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Immunofluorescent Labeling of Skeletal Muscle

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Skeletal muscle sections, 9 μm thick, were fixed in 100% methanol and washed twice in 1X phosphate-buffered saline. Sections were blocked in 0.1% Tx-100/10% fetal bovine serum/phosphate-buffered saline at room temperature for 1 hour and then incubated with primary antibodies (anti-VIA4-1, a gift from Kevin Campbell diluted at 1:20) overnight at 4°C. AlexaFluor secondary antibodies (goat anti-mouse568 1:500 + goat anti-rabbit488 1:500) were used followed by mounting with Fluoromount-G. Images were acquired on a Nikon epi-fluorescent microscope.

Meilleur K.G., Zukosky K., Medne L., Fequiere P., Powell-Hamilton N., Winder T.L., Alsaman A., El-Hattab A.W., Dastgir J., Hu Y., Donkervoort S., Golden J.A., Eagle R., Finkel R., Scavina M., Hood I.C., Rorke-Adams L.B, & Bönnemann C.G. (2014). Clinical, Pathological and Mutational Spectrum of Dystroglycanopathy Due to LARGE Mutations. Journal of neuropathology and experimental neurology, 73(5), 425-441.

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Temporal Dynamics of Neuroinflammation

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Following double immunofluorescence staining, the positively stained cells in the intramatrix zone (IMZ) and the lesion boundary zone (LBZ) were counted manually on D7, D14, D21, and D28 in three to five different fields per section of each rat brain (using an eyepiece grid covering an area of 0.0625 mm²). An individual who was blinded to the experimental design performed the counting. Vessels and blood cells were excluded. Sections were observed, and images were acquired using a Nikon epifluorescent microscope. The total cells in the LBZ were visualized by DAPI staining at 20x magnification using the Openlab software (Improvision, Cambridge, MA, USA). IBA-1 cells were visualized as fluorescent red (DL 549) and GFAP cells were visualized as fluorescent green (Alexa 488) in the LBZ at 20x magnification. The double-positive cells were visualized as yellow. For double immunofluorescence staining, the double-positive cells were also manually counted in the same way. The results were presented as the number of immunopositive cells per field.

Chen J.H., Hsu W.C., Huang K.F, & Hung C.H. (2019). Neuroprotective Effects of Collagen-Glycosaminoglycan Matrix Implantation following Surgical Brain Injury. Mediators of Inflammation, 2019, 6848943.

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