Serial bright-field and immunofluorescent images of the co-cultured mCherry-labelled hMSCs and cancer cell clusters were obtained at day 0, 3 and 7, using a Nikon Eclipse Ti-E microscope with differential interference contrast (DIC). Forty z-stacks were taken throughout the depth of the well, and extended depth of field projection picture was produced using the Nikon NIS Elements software, allowing a condensed view of the three-dimensional profile of the cancer cell clusters. On day 7, cell clusters from the 3D-TGA were extracted using the standard method, [58 (link)] and immuno-stained as previously reported, [5 ] with Trefoil Factor 3 (TFF3, Abcam), AlexFluor488 (ThermoFisher Scientific) and mounted in Prolog Gold Anti-fade containing DAPI (ThermoFisher Scientific). Immunofluorescent images of the stained clusters were obtained with the Nikon Eclipse Ti-E microscope and NIS Elements Advanced Research Software.
Nis elements advanced research software
Manufactured by National Instruments
NIS-Elements Advanced Research is an image acquisition and analysis software for microscopy applications. It provides a comprehensive set of tools for capturing, processing, and analyzing images from various types of microscopes. The software supports a wide range of imaging modalities, including fluorescence, confocal, and transmitted light microscopy.
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3 protocols using nis elements advanced research software
1
Co-culture Imaging of hMSCs and Cancer Cells
2
Quantifying E-cadherin and Blastocyst VVL-FITC
3
Quantifying E-cadherin and Blastocyst VVL-FITC
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For E-cadherin and Golgi quantitation, immunofluorescence images were captured using Nikon A1 laser scanning confocal microscopy. NIS-Elements Advanced Research software (version 4.51.00) was used to measure the intensity of E-cadherin (green) in Golgi (red). To measure the intensity of E-cadherin in the Golgi, the Region of Interest Tool was used to select the Golgi area for each cell. Then, the Perform Measurements Tool was used to determine the intensity of E-cadherin (green) in Golgi (red) for each cell. Mean intensity of E-cadherin in Golgi per cell was analyzed and plotted using GraphPad Prism7 software.
For blastocyst VVL-FITC quantitation, NIS analysis elements software was used to measure the intensity of VVL-FITC (green) in the trophectoderm. To measure the intensity of VVL, the Region of Interest Tool was used to select the trophectoderm area and the Perform Measurements Tool was used to determine the intensity. The VVL-FITC intensity for the blastocysts was analyzed and plotted using GraphPad Prism7 (version 7.0).
For blastocyst VVL-FITC quantitation, NIS analysis elements software was used to measure the intensity of VVL-FITC (green) in the trophectoderm. To measure the intensity of VVL, the Region of Interest Tool was used to select the trophectoderm area and the Perform Measurements Tool was used to determine the intensity. The VVL-FITC intensity for the blastocysts was analyzed and plotted using GraphPad Prism7 (version 7.0).
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