Fig. 15.1
(a) Vessel wall, with deposits of amyloid, in renal AL amyloidosis. Congophilia (salmon-pink color) is noted in the vessel wall, corresponding to amyloid deposits. Congo red stain, bright field. Magnification ×750. (b) Vessel wall with deposits of amyloid in renal AL amyloidosis. A distinct bright yellow-green fluorescence highlights the entire amyloid-containing area in the arteriolar wall; no equivalent to “polarization shadow” is seen (see Chap. 14). Thioflavin T stain viewed under fluorescein gate using a standard immunofluorescence microscope. Magnification ×700
Fig. 15.2
Glomerulus with extensive amyloid deposition detected by strong fluorescence with thioflavin T stain. Note, also, staining of adjacent arteriolar wall. Thioflavin T stain viewed under fluorescent light. Magnification ×750
Fig. 15.3
(a) Eosinophilic, amorphous (hyaline) material corresponding to areas with amyloid deposition noted in the three renal compartments: glomerulus, interstitium, and extraglomerular vessels. Hematoxylin and eosin stain, original magnification ×350. (b) Distinct fluorescence highlighting amyloid deposits in the three renal compartments: glomerulus, interstitium, and extraglomerular vessels. Thioflavin T stain, original magnification ×350
Thioflavin T Stain for Amyloid
A working solution of 0.5 % of thioflavin T in 0.1 N HCl (mixed well and filtered before use, aliquoted, and stored frozen as needed) is used. The sections are deparaffinized and rehydrated. Using a hydrophobic marker, the specimen section and an adjacent positive control section are encircled on the top of the slides. A drop of the working solution of thioflavin T is placed on the slides, which are subsequently kept in a humidity chamber for 15 min. The slides are rinsed in deionized water (for 5 min) and kept in deionized water while coverslipping with Aquamount. The frozen sections are rinsed two times in PBS (for 10 min each) prior to incubation with thioflavin T and, thereafter, rinsed sequentially in PBS and deionized water (for 5 min each) and coverslipped. Thioflavin stain is viewed under a fluorescein gate using a standard immunofluorescence microscope. Thioflavin T fluoresces only when bound to amyloid fibrils, where it shows a bright yellow-green fluorescence. In contrast, in the absence of amyloid deposits, the dye fluoresces faintly.