Double dark-field staining method, stained specimen, and use of enzymatic tracers in double dark-field staining

摘要

A method is presented for a dark-field immunohistochemical double staining approach. A preferred embodiment combines the use of an immuno-alkaline phosphatase technique and the immunogold-silver technique, visualized with fluorescence- and epipolarization illumination, respectively. In this alkaline phosphatase activity revealing technique, the reaction product shows an intense red fluorescence with a rhodamine filter set and no signal with epipolarization illumination. The silver precipitate does not exhibit any signal using the rhodamine filter set. This allows separate observation and photographic recording of two antigens in one tissue section; an objective that cannot be achieved with conventional immunoenzyme double staining methods. The double dark-field approach presented is compatible with different immunoenzyme double staining protocols.