texas-red and Amyloidosis

The tissue diagnosis of amyloidosis is traditionally suggested by hematoxylin-eosin stain and confirmed by Congo red stain, both examined by routine light microscopy. Both false-positive and false-negative congophilia are well documented, limiting the sensitivity and specificity of the Congo red stain for the diagnosis of amyloidosis. Examination of Congo red-stained tissue by Texas Red-filtered fluorescence microscopy (TRFM) is known to enhance the amyloid-specific congophilia, thus increasing the diagnostic sensitivity.. To determine whether TRFM can mitigate the false positivity and thus improve the diagnostic specificity of the Congo red stain.. Ninety-two tissue samples were categorized into 3 groups. Group I included 15 samples with tissue deposition of amyloid. Group II consisted of 63 samples in which amorphous eosinophilic structures reminiscent of amyloid were seen on hematoxylin-eosin-stained tissue sections. Group III included 14 samples in which amyloid and amyloid-like tissue were seen side by side. The final diagnosis of presence or absence of amyloidosis in each case was established by clinicopathologic correlation. The congophilic areas in each case were identified by light microscopy. The same areas were then examined by TRFM.. TRFM enhanced congophilia, confirming the diagnosis of amyloidosis in all group I cases. Enhancement was not seen in 52 of the 63 group II cases. For group III cases, TRFM enhanced the amyloid-specific congophilia, but not the nonspecific congophilia, in all cases.. TRFM increases the diagnostic yield and specificity of Congo red-stained tissue sections for detection of amyloid.

Topics: Amyloid; Amyloidogenic Proteins; Amyloidosis; Congo Red; Eosine Yellowish-(YS); Hematoxylin; Humans; Microscopy, Fluorescence; Staining and Labeling

Murine serum amyloid A1.1 (SAA1.1) has been conjugated with the fluorophore Texas Red (TxR), and its interaction with peritoneal macrophages has been visualized by scanning confocal microscopy. Binding of TxR-SAA to cell surfaces was inhibited by an excess of unlabelled SAA indicating the involvement of saturable receptors. Internalized TxR-SAA was seen initially as small punctate signals which in some cells evolved into a fine fluorescent network, a pattern typical of tubular endosomes. Colocalization of TxR-SAA with Cy5-labelled low density lipoprotein (LDL) but not with Oregon Green-labelled transferrin suggested that SAA trafficked through endosomes and lysosomes for degradation rather than through recycling compartments. Consistent with this catabolic pathway, macrophages loaded with TxR-SAA lost fluorescence within several days after being shifted to a fluorophore-free medium. In sharp contrast to this, cells maintained under amyloid-forming conditions, i.e. in the presence of unlabelled SAA and amyloid-enhancing factor (AEF) before and after treatment with TxR-SAA, remained brightly fluorescent over the course of 5 days. Immunocytochemistry verified the accumulation of SAA within macrophages. These findings support the hypothesis that a decreased catabolism of internalized SAA plays a role in AA amyloid pathogenesis.

Topics: Amyloidosis; Animals; Apolipoproteins; Biological Transport, Active; Fluorescent Dyes; Immunohistochemistry; In Vitro Techniques; Macrophages, Peritoneal; Mice; Mice, Inbred C57BL; Microscopy, Confocal; Protein Binding; Serum Amyloid A Protein; Xanthenes