DIGOXIGENYLATED PRIMARY ANTIBODIES FOR SENSITIVE DUAL-PEROXIDASE LABELING OF NEURAL MARKERS

This study extends the application of the digoxigenin-anti-digoxigenin (DIG) technique to immunocytochemistry by using digoxigenin-tagged pr imary antibodies. Certain features of this technique when applied to n on-radioactive in situ hybridizaton, such as the absence of endogeneou s digoxigenin immunoreactivity in animal tissues, seem to be advantage ous also for its application to immunocytochemistry. Thus, the present work is focused on dual-peroxidase staining experiments based on digo xigenylated antibodies directed against glial fibrillary acidic protei n, parvalbumin, and calbindin, in a straightforward combination with c onventional cytochemical methods. The protocols include the concomitan t detection of two antigens, for which only primary antibodies from on e animal species are available, with differently haptenized antibodies (e.g., biotinylated anticalbindin and digoxigenylated anti-parvalbumi n). The versatility of the DIG technique is exemplified by the combina tion of lectin and immunocytochemical procedures for the detection of astrocytes and microglia, and the simultaneous visualization of perine uronal nets and parvalbumin-containing neurons in the rat brain.