ACVP-03: Novel CD4+ T Cell Specific Immunohistochemistry Detection and Analysis Utilizing Masking of Not-T Cell CD4 in Fixed Tissues from Virally Infected and Uninfected Specimens

The Tissue Analysis Core (TAC) within the AIDS and Cancer Virus Program will process, embed, and perform microtomy on fixed tissue samples presented in ethanol. CD4 (DAB) and CD68/CD163 (FastRed) double immunohistochemistry will be performed, in which the CD68/CD163 signal “masks” the low CD4 expression found on macrophage/dendritic cells. To ensure consistency among assay batches, we have automated a major portion of the double immunohistochemistry assay using the IntelliPATH autostainer. A high-resolution digital image of each double immunohistochemistry stained sample will be generated using the Aperio ScanScope CS System. Regions of interest (ROI; 20–30) of a defined size will be highlighted on each digital image using the Aperio ImageScope software regions tool, and high-resolution images of ROIs will be generated using a custom-written software program created by the Imaging and Visualization Group that automatically extracts ROIs from the digital slide annotation layer. Finally, under the guidance of a subject matter expert, the TAC will provide comprehensive quantitative image analysis using a custom series of unique actions in Photoshop with Fovea tools to determine the percent area in each high-resolution image that is occupied by CD4+ T cells. Relevant data will be provided electronically, and a report with interpretation/conclusions will be supplied. Sufficient basic consultation with a subject matter expert is included with the service.