Vs120 s6 slide scanner

AB-PAS was performed to assess the presence of mucin using Tissue-Tek Prisma. In brief, the organoids were stained with an alcian blue solution with a pH of 2.5 for 8 min, periodic acid for 10 min, followed by a 3 min rinse with water after each staining. Finally, the organoids were stained with schiff reagent for 15 min, rinsed with tap water for 5 min and stained with hematoxylin for 30 sec. The sections were cleared using xylene for 2 min. AB-PAS-stained samples were scanned with a 40x objective, using the VS120 S6 Slide scanner (Olympus, Japan), and the images were taken with the color camera, pike F-505 (Allied Vision). The images were processed using the ImageJ software.

The mice were perfused with 20 ml of PBS, followed by 40 ml PBS containing 4% paraformaldehyde (PFA) using a mini-pump with variable flow (United States Plastic Corp., US). The perfused brains were fixed in 4% PFA solution for 24 h and then soaked in a 30% sucrose-PBS solution for another 24 h at 4°C. Then the brains were frozen in dry ice and coronally sectioned at 30 μm thickness using a microtome (Leica SM2010R, Germany). One out of every three consecutive sections were mounted, coverslipped, and imaged. To examine the expression of amyloid-beta, two sections from each of five brains from each mouse group (10 sections in total for each mouse group) were stained with primary antibody 6E10 (Biolegend, Mouse, #803002,1:500 dilution, US), followed by Cy3 conjugated donkey anti-mouse secondary antibody (Jackson ImmunoResearch, 1:200 dilution, US). All sections were counterstained with 10 μM DAPI. Images were acquired by an automated fluorescent slide scanner (Olympus VS120-S6 slide scanner, Japan) using a 10× magnification objective. For higher resolution imaging, selected slices were imaged using a confocal microscope (Olympus FLUOVIEW FV3000, Japan) with a 40× magnification objective.

H&E staining of FFPE samples was performed to assess the histological features of the organoids. H&E staining was done according to standard protocols as previously described by Li et al., (Li et al., 2018 (link)). H&E-stained samples were scanned with a 40x objective, using the VS120 S6 Slide scanner (Olympus, Japan) and the images were taken with the color camera, pike F-505 (Allied Vision). The images were processed using the ImageJ software.