Prior to digestion, a portion of the RCC sample and normal adjacent tissue were fixed in 10% neutral buffered formalin at room temperature for 48 hours. Tissues were then dehydrated in 70% ethanol for 24 hours. The samples were then embedded in paraffin and processed by the KUMC BRCF staff using a Tissue-Tek Processor (Sakura). Microtomy was performed on a Leica RM2255 Microtome (Leica Biosystems). Automated staining for pan-cytokeratin and the paired box 8 protein (PAX-8) was performed using an intelliPATH instrument (Biocare). Immunocytochemistry was performed using the same protocol with cell pellets of the primary TCLs established from the tumor sample. Images were acquired using MetaMorph Microscopy Image Analysis Software (Molecular Devices) on a Nikon Eclipse 80i Microscope.
Metamorph microscopy image analysis software
Manufactured by Molecular Devices
Sourced in United States
MetaMorph® Microscopy Image Analysis Software is a comprehensive image analysis platform designed for researchers. It provides tools for acquiring, processing, and analyzing digital images from a variety of microscopy techniques. The software offers a wide range of image analysis capabilities, including image segmentation, object detection, and quantitative measurements.
Automatically generated - may contain errors
Lab products found in correlation
Rm2255 microtome, by Leica (1 mentions)
Eclipse 80i microscope, by Nikon (1 mentions)
Anti mouse cd68, by Bio-Rad (1 mentions)
Anti mouse mmp 9 antibody, by R&D Systems (1 mentions)
Phalloidin tritc, by Merck Group (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Merck Group (1 mentions)
F0054, by Agilent Technologies (1 mentions)
A0082, by Agilent Technologies (1 mentions)
X0936, by Agilent Technologies (1 mentions)
Masson s trichrome, by Merck Group (1 mentions)
Anti α sma, by Merck Group (1 mentions)
Anti f4 80, by Abcam (1 mentions)
4 protocols using metamorph microscopy image analysis software
1
Immunohistochemical Analysis of RCC Samples
2
Immunohistochemical Analysis of Aortic Lesions
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Mouse aortic sinuses were serially cut at 5 μm as previously described [20 (link),21 (link)]. The obtained sections were fixed in acetone and immunostained with specific anti-mouse MMP-9 antibody (R&D Systems, Minneapolis, MN, USA), anti-mouse CD68 (Serotec, Puchheim, Germany). All sections were counter-stained with Mayer’s hemalum solution and rinsed in distilled water. Quantification was performed using the MetaMorph® Microscopy Image Analysis Software from Molecular Devices or Definiens Developer 2.7 software (Definiens Inc., Carlsbad, CA, USA) for biomarker and morphological tissue profiling. The results were expressed as a stained area in lesion versus total lesion area.
3
Immunofluorescent Characterization of BOECs
4
Kidney Fibrosis Analysis via Microscopy
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To analyze kidney brosis, kidney sections were stained with Sirius red, Masson's trichrome, and anti-α-SMA (Sigma) and examined by light microscopy. Positive Sirius red, Masson's trichrome, and and anti-α-SMA (Sigma) staining areas were evaluated relative to the unit area and expressed as a percentage per unit area using MetaMorph microscopy image analysis software (Molecular Devices, Sunnyvale, CA, USA). Macrophage immunohistochemistry was performed using anti-F4/80 (Abcam) antibodies and F4/80-positive cells quanti ed as the number of cells per high power eld [24] . Microscopy assessment was carried out in a blinded manner, with 20 randomly selected elds from each slide section examined at ×400 magni cation. To trace MPs in vivo, freshly isolated MPs were labeled with red uorescent CellTracker™ and detected within the peritubular interstitium of kidney sections by immuno uorescence microscopy.
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