FLUORESCENCE OVERLAY ANTIGEN MAPPING OF THE EPIDERMAL BASEMENT-MEMBRANE ZONE .1. GEOMETRIC ERRORS

To identify in tissue sections the relative positions of antigen distr ibutions close to the resolving power of the microscope, we have devel oped the fluorescence overlay antigen mapping (FOAM) procedure. As thi s technique makes high demands on the geometric fidelity of the overla y image, it is essential to recognize geometric errors resulting from optical imperfections. This applies in particular to the image shift d ifference (ISD) that may routinely occur during fluorescence overlay. We describe here procedures for assessment and mechanical correction o f the ISD in tissue sections. Furthermore, we describe an alignment ve rification test to assess the accuracy of the ISD correction procedure , using collagen Type VII as the geometric verification marker. These procedures should enable reliable evaluation of relative antigen distr ibutions in tissue sections using photomicrographic multicolor fluores cence overlay. Further details of the FOAM technique, such as color fi delity and its utility for diagnostic and research purposes, will be p ublished separately.