We used Vectra 2.0 multispectral imaging system (Perkin Elmer) to image the multiplexed IHC slides (Fig. 2 ). This is an automated and multimodal imaging system that not only captures high-quality images of the entire tissue section or within the regions of interest but also allows to study the morphometric and biochemical organization of an intact tissue section both qualitatively and quantitatively. Its multiplexing capability can separate up to eight markers/fluorophores within the field of view in a single tissue section. For more information on how to use the latest version of the microscope, please find the user manual from the manufacturer in this link: https://resources.perkinelmer.com/corporate/content/lst_software_downloads/vectra-user-manual-3-0-3.pdf .
The following steps represent an overview of using the instrument for imaging of the IHC multiplexed TMAs. Note that these steps are specific for the Vectra 2.0 multispectral imaging system (Perkin Elmer) installed at The Ohio State University Comprehensive Cancer Center:
If your experimental goal is to detect only one antigen with a single fluorophore (e.g., if you are interested to look at the membranous expression of ENT1 in your samples tagged with the Opal 570 fluorophore) and you would like to capture the images from multiple random fields, then the following steps would apply:
The following steps represent an overview of using the instrument for imaging of the IHC multiplexed TMAs. Note that these steps are specific for the Vectra 2.0 multispectral imaging system (Perkin Elmer) installed at The Ohio State University Comprehensive Cancer Center:
If your experimental goal is to detect only one antigen with a single fluorophore (e.g., if you are interested to look at the membranous expression of ENT1 in your samples tagged with the Opal 570 fluorophore) and you would like to capture the images from multiple random fields, then the following steps would apply: