In efforts to better understand the cell biology of fungi and their pathogenic interactions with plants, we have pursued the use of cryo-preparative techniques for specimen preservation and immunocytochemistry at the light and electron microscope level. The chemically diverse composition of plant and fungal walls has proven very challenging for immunofluorescence using confocal microscopy. Typically, to allow probe access in these organisms, immunolabeling has required sectioning or enzymatic digestion of cell walls. The ideal method would preserve the spatial fidelity and antigenicity of the target molecule as well as maintain the integrity of the surrounding tissue. Our early work using methacrylate de-embedment, as described for sectioned plant material, was very effective for freeze-substituted fungi. Further improvements, using a combination of slow freezing and freeze-substitution followed by methacrylate de-embedment, permitted whole mount fluorescence antibody labeling of a variety of phylogenetically diverse fungi without enzymatic digestion or sectioning (Fig.l).