Anti transthyretin

Following postfixation of the brain samples in formalin, 10‐μm thick coronal sections were cut using a cryostat (Leica Microsystems LM3050S). Brain sections revealing choroid plexus were used for immunohistochemical staining as previously described.15 Briefly, these brain sections were incubated overnight at 4°C with the following primary antibodies: anti‐IRP1 (1:200, Abcam), anti‐IRP2 (1:100, Abcam), anti‐NCBE (1:50, Origene), and antitransthyretin (1:50, Abcam). Sections were then washed and incubated with the appropriate secondary antibodies (1:100, Jackson ImmunoResearch Labs) for 2 hours at room temperature. The tissue slides were then stained with 4’,6‐diamidino‐2‐phenylindole (Vector Laboratories), fixed in paramount, and visualized using a fluorescence microscope (Olympus BX51, Olympus Corporation). For histological, volumetric analysis, 16‐μm thick coronal brain sections were cut at 2.5 mm, 1.2 mm, 0.7 mm rostral, and 2.9 mm caudal of the bregma. These brain slices were stained with Nissl and morphometrically analyzed using ImageJ 4.0 (Media Cybernetics) assisted delineation of brain structures, as previously described.16 The ventricle volume was calculated as average ventricular areas on each slide multiplied by the depth of the cerebroventricular system.