COMPUTER-AIDED IMAGE-ANALYSIS APPLIED TO IMMUNOGOLD-SILVER STAINING -EVALUATION OF PROLIFERATING CELL NUCLEAR ANTIGEN (PCNA)-REACTIVE SITES IN PARAFFIN SECTIONS

Feasibility of the combination of the immunogold-silver staining metho d (IGSS) and computer-aided image analysis was assessed for the detect ion of antigen in an immunostained, paraffin-embedded section. Using l ow-temperature IGSS, we stained a specimen of human oral squamous cell carcinoma with a monoclonal antibody, PC 10, against a proliferating cell nuclear antigen (PCNA/cyclin), and the section was analyzed by AC AS 570 interactive laser cytometry. The PCNA-positive cells, exhibitin g a heteromorphic texture, were contrasted by the dark staining of the ir nuclei, but showed heterogeneity in staining intensity from cell to cell. Using a conventional microscope light source rather than a lase r, and by employing the COMPLEMENT DATA program (which permits inversi on of the data values) installed in the ACAS 570 software system, we w ere able to obtain a 'complemental image' which replicated the real im munohisto-morphology. Approximately 30-35 cells from three different a reas in the same section were selected by DEFINE CELL and MARK AREA pr ograms, and quantitative image analysis was performed in terms of cell integrated value, area, perimeter, and shape factor indicated in hist ogram form. The combined utilization of IGSS with computer-aided image analysis was demonstrated to offer a crucial advantage for the quanti tative assessment of immunostained sections.