Ventana immunostainer

Immunohistochemistry for neuropathological evaluation was performed on small paraffin-embedded sections from right brain hemisphere obtained from the same cases mentioned above (AβPParc1, AβPParc2, and PSEN1DE9) (For AβPParc1 6 μm thick sections on no coated slides were used; for AβPParc2 7 μm thick sections on coated slides were used and for PSEN1DE9 7 μm thick sections on superfrost slides were used).
A routine deparaffinization protocol was used. AT8 (Phospho-Tau, Ser202, Thr205) monoclonal antibody from Thermofisher, amyloid beta 1–42 (antiamyloid β42 antibody, clone G2-11 from Merck Millipore), and amyloid beta 1–40 (antiamyloid β40 antibody, clone G2-10 from Merck milipore) antibodies were used as follows: AT8 dilution 1:2500 stained in Roche Ventana immunostainer; amyloid beta 1–42 dilution 1:750 and amyloid beta 1–40 dilution 1:500 stained in Roche Ventana immunostainer but with 10 min in formic acid first after deparaffination.

Hematoxylin and eosin staining was performed on 5 μm sections from the paraffin-embedded tissue blocks for conventional light microscopy analysis. For CD68 staining, the liver sections were de-paraffinised and immunostained with rabbit anti-CD68 antibody (Abcam) using the automated Ventana immunostainer, according to manufacturer's recommendation. For CD3 staining, the liver sections were de-paraffinised and immunostained with rabbit monoclonal anti-CD3 antibody (Ventana Medical Systems, Inc) using the automated Ventana immunostainer, according to manufacturer's recommendation. For Sirius Red staining, liver sections were de-parrinised and stained using standard protocol.