Transmission electron micrographs (TEM) and immunogold labeling of experimental and fossil tissues with antiserum to feather keratin.

Ab-ag complexes are demonstrated by electron-opaque gold beads attached to the secondary antibody. A-C) Localization of ab-ag complexes to keratinous tissues in the control feather. Melanosomes can be seen in A, B), but virtually no gold beads localize to these structures, and remain localized only to the filamentous matrix, supporting antibody specificity. C) gold beads are specifically associated with electron-lucent filaments against a darker background. D-F show the same immunoassay results on the condition 1 feathers. Melanosomes can be seen, but they are less electron dense than in A), and most exhibit hollow cores, possibly indicating initial degradation. Again, gold beads, reflecting the location of ab-ag complexes, are localized to the keratinous matrix interspersed between melanosomes, although these are reduced in density from the control feathers. F) Edge of a melanosome (arrow); no binding of the small gold beads is observed on the melanosome, but is localized to the matrix surrounding the melanosome. G-I) Immunolabelling on a small region of the condition 2 feather; although no melanosomes are seen in feathers from this condition, the keratinous matrix remains. Binding of antibodies is sparse, but is specific and highly localized to remnants of electron-lucent filaments (H, I). J-L) Localization of gold beads to Yellowstone coot feathers under low (J) and higher (K, L) magnification. No melanosomes are visible in TEM, but gold beads are strictly localized to regions of small electron lucent filaments, as in the other conditions presented here.