A simple and cost-effective technique to quench autofluorescence in formalin-fixed paraffin-embedded cnidarian tissues

Immunofluorescence is a first-choice technique for the study of neuroanatomy and, in general, of very fine structures. A stumbling block to the application of immunofluorescence is autofluorescence in tissues. This problem has different origins, as it can be related to the structures that are being examined, the fixation protocol, and the general condition of the organism which has been studied. Several techniques are available to help reduce autofluorescence in vertebrate tissues, particularly in mammalian tissues, and invertebrate tissues alike. The study of animals from different phyla generates additional problems regarding autofluorescence because of the presence of peculiar substances (e.g., chitin). In the present work, we present a simple technique to quench autofluorescence in sections of paraformaldehyde-fixed, paraffin-embedded cnidarians of three species: Paramuricea clavata, Eunicella cavolinii and Savalia savaglia. The technique is based on the histological dye Sudan Black, which is already used in mammalian tissue samples; however, to our knowledge, this work represents the first use of this method in cnidarians. This method is perfectly compatible with immunofluorescence, as it does not appear to interfere with fluorochromes that are conjugated to secondary antibodies and appears to reduce background autofluorescence.