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The application of the Phoroscope electrophoresis densitometer for the objective evaluation of immunoperoxidase-stained tissue sections at the light microscope level is described. The instrument directs a beam of light through the specimen (Kodachrome photomicrograph slides) and projects an absorbance pattern (as peaks and valleys) on an oscilloscope screen. The height of a peak is proportional to the optical density of a stained area, while the width of the peak is proportional to the diameter of an area. The utility of the densitometer was tested in sections of rat pituitary gland immunostained with NIAMDD antiserum against rat prolactin to reveal prolactin cells and sections of rat ventral prostate pretreated with NIAMDD rat prolactin and then immunostained to reveal Golgi region-localized prolactin binding sites in epithelial cells. The densitometer quantitated (by relative absorbance) the changes in immunostaining in serial sections of rat pituitary gland reacted with different dilutions of prolactin antiserum and in serial sections of rat ventral prostate pretreated with different concentrations of rat prolactin. In addition, the densitometric pattern provided information regarding the frequency and pattern of immunostaining within a particular microscopic field. Potential uses of an electrophoresis densitometer for the quantitation and standardization of immunohistochemistry are discussed.

Evaluation of immunoperoxidase-stained tissue sections with an electrophoresis densitometer.