Imagej

Manufactured by Nikon

Sourced in Japan, United States

ImageJ is a free, open-source image processing software developed by the National Institutes of Health (NIH). It is designed to analyze, process, and manipulate digital images and supports a wide range of image formats. ImageJ provides a core set of functions for tasks such as image enhancement, measurement, and analysis.

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Lab products found in correlation

Nis element, by Nikon (10 mentions) Nis elements software, by Nikon (4 mentions) Nis elements ar, by Nikon (3 mentions) A1r confocal microscope, by Nikon (3 mentions) Axio observer z1, by Zeiss (3 mentions) Lsm 710 confocal microscope, by Zeiss (3 mentions) Smz 18 zoom stereo microscope, by Nikon (2 mentions) Eclipse 80i, by Nikon (2 mentions) Prism 8, by GraphPad (2 mentions) Pronase, by Merck Group (2 mentions) Nis elements advanced research, by Nikon (2 mentions) Eclipse ti, by Nikon (2 mentions) Ez c1, by Nikon (2 mentions) Fibrinogen, by Merck Group (1 mentions) Thrombin, by Merck Group (1 mentions) Evos cell imaging system, by Thermo Fisher Scientific (1 mentions) Cytodex 3 microcarrier beads, by Merck Group (1 mentions) Aprotinin, by Merck Group (1 mentions) Matlab, by MathWorks (1 mentions) Origin 9, by OriginLab (1 mentions)

73 protocols using imagej

Quantifying p-rpS6 and Ki67 Expressions

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To study the expressions of p-rpS6 and Ki67, ICC-IF was used. Primary antibodies p-rpS6 (cell signalling, Cat# 2211, 1:500 dilution) and Ki67 (Santa Cruz, Cat# sc23900, 1:200 dilution) were incubated with the cells on slides overnight at 4 °C. Stained cells were visualized with a fluorescent microscope (Nikon) and fluorescence intensities were quantified by ImageJ (ImageJ.nih.gov">ImageJ.nih.gov).

Yang-Kolodji G., Mumenthaler S.M., Mehta A., Ji L, & Tripathy D. (2015). Phosphorylated ribosomal S6 (p-rpS6) as a post-treatment indicator of HER2 signalling targeted drug resistance. Biomarkers : biochemical indicators of exposure, response, and susceptibility to chemicals, 20(5), 313-322.

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Quantifying Cardiomyocyte Proliferation

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Images were acquired utilizing the Nikon A1R confocal microscope using ×10, ×40 and ×60 objectives (Nikon). Raw imaging data were processed using Nikon Software or ImageJ. Ki-67 stained hearts of Cdh5CreER lineage were analyzed using ImageJ, nuclei were assessed on tdTomato⁺ and Ki-67⁺ signal and quantified accordingly.

Peisker F., Halder M., Nagai J., Ziegler S., Kaesler N., Hoeft K., Li R., Bindels E.M., Kuppe C., Moellmann J., Lehrke M., Stoppe C., Schaub M.T., Schneider R.K., Costa I, & Kramann R. (2022). Mapping the cardiac vascular niche in heart failure. Nature Communications, 13, 3027.

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Imaging Zebrafish Embryos at 2 dpf

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For gross imaging of 2 dpf embryos, embryos were dechorionated at described stages with incubation in 0.03% pronase (Sigma-Aldrich, P5147) for 6 min at 1 dpf. Embryos and adults were anesthetized using 0.4% tricaine. For light and immunofluorescence imaging, embryos were mounted in 1% low-melting-point agarose in a 60×15 mm Petri dish. Gross images and images with DASPEI or AR staining were taken on a Nikon SMZ-18 Zoom Stereo Microscope. For confocal imaging, embryos were mounted on a glass-coverslip-bottomed dish and imaged using a Nikon A1 confocal with a 60× oil-immersion objective. For quantification, all images were acquired at the same magnification, laser power, exposure time and gain. After each embryo was imaged, embryos were removed from the agarose to generate genomic DNA for genotyping. Further figure processing and analysis was performed using Nikon NIS Element and ImageJ.

Wang J., Thomas H.R., Thompson R.G., Waldrep S.C., Fogerty J., Song P., Li Z., Ma Y., Santra P., Hoover J.D., Yeo N.C., Drummond I.A., Yoder B.K., Amack J.D., Perkins B, & Parant J.M. (2022). Variable phenotypes and penetrance between and within different zebrafish ciliary transition zone mutants. Disease Models & Mechanisms, 15(12), dmm049568.

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3D Fibrin Gel Assay for Angiogenic Potential

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3D fibrin gel assay was performed as previously described^{25 (link)}. Briefly, Cytodex® 3 microcarrier beads (Sigma) were incubated with HUVEC, HUVEC-TIE2-WT or HUVEC-TIE2-L914F at a concentration of 400 cells/ bead for 4 h at 37 °C. The following day, coated beads were resuspended in 2 mg/mL of fibrinogen (Sigma) solution containing 0.15 U/mL of aprotinin (Sigma) at a concentration of 500 beads/ml. Then 0.625 U/mL of thrombin (Sigma) and 0.5 ml beads/fibrinogen suspension were added per well of a 24-well plate and incubated at 37 °C to allow fibrin clotting. The gels were overlaid with human lung fibroblasts at 2 × 10⁴ cells/well and medium was replaced every other day. Where indicated fibroblasts were omitted and EGM2/10% FBS medium substituted with EBM2 (Endothelial basal medium) which does not contain growth factors. In assays conduced with pericytes the Cytodex® beads were seeded with EC (HUVEC or HUVEC-TIE2-L914F) and pericytes at a 10:1 ratio. Images were acquired with the Nikon A1R LUN-V Inverted Microscope and EVOS cell imaging system (Invitrogen) and analyzed with Nikon NIS-Elements and ImageJ.

Cai Y., Schrenk S., Goines J., Davis G.E, & Boscolo E. (2019). Constitutive Active Mutant TIE2 Induces Enlarged Vascular Lumen Formation with Loss of Apico-basal Polarity and Pericyte Recruitment. Scientific Reports, 9, 12352.

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3D Confocal Image Reconstruction

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3D reconstruction of the confocal image was done with Nikon confocal imaging program and ImageJ. Z-stack projection is done with summation or max intensity projection depending on the data requirement. The raw image was 16-bit, but the processed images were stored in 8-bit or RGB for compatibility with documentation.

Jo Y., Yim D., Park C.E., Yong I., Lee J., Cho W., Ahn K.H., Yang C., Chang J.B., Park Y.G., Kim T.S., Kim T, & Kim P. (2023). Bi-directional crosstalk between cells and extracellular matrix leads to network morphogenesis in multi-layered tissues. Research Square.

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Mouse Model of Lung Tumor Initiation

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All animal studies were IACUC approved. The Smad4 conditional knockout allele, lox-stop-lox-Kras^G12D conditional knock-in allele, and K5Cre*PR transgene, and K14CrePR transgene have been previously described (26 (link), 27 (link), 46 (link)-48 (link)). Lung tumors were initiated with 500μg tracheal RU486 at 4-6 weeks of age. Mice were euthanized between 12-18 months of age or if they lost >15% of their body weight; tumors were measured, enumerated, collected, then classified according to consensus criteria (49 (link)). PCR for recombinant Kras and Smad4 alleles was performed as previously described (50 , 51 (link)). Smad4 and pH2AX immunostaining was performed as described above. Tumor cell size was analyzed by counting the number of cells in a 40X field from H&E stained sections from at least 5 animals of each genotype. All images were acquired on a Nikon Eclipse 80i microscope with Nikon Elements Software and analyzed with ImageJ (rsbweb.nih.gov). Differences between groups were compared with by one-way ANOVA, unpaired t test, or Fisher's exact test as appropriate.

Haeger S.M., Thompson J.J., Kalra S., Cleaver T.G., Merrick D., Wang X.J, & Malkoski S.P. (2015). Smad4 loss promotes lung cancer formation but increases sensitivity to DNA topoisomerase inhibitors. Oncogene, 35(5), 577-586.

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Quantitative Fluorescence Analysis

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Statistical analysis was conducted using Graphpad Prism 8.0 software. Linescan analyses and other fluorescence intensity measurements were conducted using Nikon Elements or ImageJ software, following background subtraction. A paired t-test was used to compare two conditions, while multiple conditions were compared using a two-way ANOVA followed by Tukey’s multiple comparison test or Dunnett’s test. See figure legends for details.

Shankar R., Lettman M.M., Whisler W., Frankel E.B, & Audhya A. (2022). The ESCRT machinery directs quality control over inner nuclear membrane architecture. Cell reports, 38(3), 110263.

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Time series image analysis protocol

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Time series images were analyzed using Nikon NIS and ImageJ with a stack stablizing plugin (http://www.cs.cmu.edu/~kangli/code/Image_Stabilizer.html). A mean background value obtained from regions away from the pHluorin was subtracted in order to correct for fluorescence intensity (F). For each pixel, we calculated the normalized change in fluorescence intensity (ΔF/F₀), where F₀ is the baseline fluorescence averaged from five frames obtained prior to light stimulation. ΔF/F₀ over time was further processed using Origin 9.1 (OriginLab). ΔF/F₀ images were processed with MATLAB (MathWorks) using custom-written scripts in order to produce pseudocolor images which is provided as Source code 1.

Wu L., Dong A., Dong L., Wang S.Q, & Li Y. (2019). PARIS, an optogenetic method for functionally mapping gap junctions. eLife, 8, e43366.

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Meiosis Imaging and Analysis Protocol

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Timing: 1–5 days depending on the number of plates imaged

This step describes how to analyze the cells after imaging.

40.

Analyze the acquired images for the phenotype of interest using a software system compatible with the images taken on the microscope (like Nikon NIS Elements or ImageJ).

Note: For our analysis we used Nikon NIS Elements. We opened each image set and made a maximum projection image from the z-stacks, overlaying GFP and mCherry fluorescence. We used the DIC image to ensure that we were analyzing cells in a monolayer and to see whether cells had budded.

Note: In our screen, we were looking for cells that stayed committed to meiosis. Therefore, we scanned 10-fields per-well of the 96-well plates and counted the number of cells that had remained in meiosis, and those that had formed buds.

Gavade J.N, & Lacefield S. (2022). High-throughput genetic screening of meiotic commitment using fluorescence microscopy in Saccharomyces cerevisiae. STAR Protocols, 3(4), 101797.

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Transwell Invasion Assay for HTR-8/SVneo Cells

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A serum-free suspension of cultured cells was loaded onto the Matrigel™-coated upper parts of Transwells (pore size of 8 μm; 5 × 10⁴cells/insert; Costar; Corning) in triplicate. The lower parts of the Transwells contained culture media containing 10% fetal bovine serum. The assembled 24-well plates were incubated for 18 h in a humidified atmosphere of 5% CO₂. Invading HTR-8/SVneo cells that had entered the lower surface of the filter membrane were stained with Wright-Giemsa Stain Kit (Nanjing JianCheng Technology co., LTD, D010-1-2, China), photographed using a light microscope (Eclipse; Nikon; Tokyo, Japan), and counted by using ImageJ (San Diego, CA, USA), whereas non-invading HTR-8/SVneo cells were removed carefully with a cotton swab. Each assay was carried out in triplicate wells.

Chen J., Qiu M., Huang Z., Chen J., Zhou C., Han F., Qu Y., Wang S., Zhuang J, & Li X. (2020). Nicotine Suppresses the Invasiveness of Human Trophoblasts by Downregulation of CXCL12 Expression through the Alpha-7 Subunit of the Nicotinic Acetylcholine Receptor. Reproductive Sciences, 27(3), 916-924.

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