Quantitative Immunocytochemistry by Digital Image Analysis: Application to Toxicologic Pathology

Abstract

Recent advances in immunocytochemical techniques allow the localization of specific antigens in tissue sections. The work reported here attempts to evaluate the application of antibody-labeled, disease-related protein, followed by microscopy and computerized image analysis. Using an experimental anti-tumor, polyclonal antibody (anti-oncomodulin) as a model, various tissues were prepared for light microscope immunocytochemistry. Sections were incubated with primary antibody, then biotinylated secondary antibody. This was followed by incubation with avidin-biotin-peroxidase (ABC method). Marker was visualized by the presence of precipitated diaminobenzidine. Samples were evaluated using a Zeiss/Kontron IBAS I & II semi-automatic digital image analysis system. Statistical analyses were performed on output data. Results demonstrated the localization and determined optical density of immunolabel. Statistical comparisons showed significant differences between control and experimental sections. The practical application of these combined techniques provides the toxicologic pathologist with a powerful tool for accurate and objective determination of the location and relative amount of selected proteins in normal and abnormal tissues.