Colocalization of ubiquitin and serum amyloid A and ubiquitin-bound AA in the endosomes-lysosomes: A double immunogold electron microscopic study

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Abstract
Alveolar hydatid cyst infection in mice induces multiple organ AA deposition at 1 week postinfection (p.i.) and increased expression of ubiquitin (UB) in both free and fixed monocytoid cells. We examined the distribution of UB, serum amyloid A and AA at 1 week p.i. in splenic perifollicular reticuloendothelial (SP-RE) cells using double immunogold electron microscopy. One side of the grid bearing the ultrathin sections was immunostained with chicken anti-bovine UB IgG-rabbit anti-chicken IgG conjugated to 6 nm colloidal gold and the other side with rabbit anti-mouse AA IgG-goat anti-rabbit IgG conjugated to 15 nm colloidal gold. Two major patterns of gold labeling were observed. The 6 nm particles, indicative of UB, labeled cytoplasmic and various subcellular orgenelles in SP-RE cells. Co-deposition of 6 and 15 nm particles, the latter indicative of AA epitope reactivity, was restricted to dense endosomes-lysosomes (EL), intravesicular “stumpy” and extracellular slender AA fibrils. Sections reacted with the adsorbed primary antibodies were not immunoreactive. The results show that UB and SAA colocalize in the EL and UB binds to both intravesicular and extracellular AA. The presence of UB label at each of these sites may suggest a physiological role for UB in SAA processing.