Anti h2a z

Embryonic small intestines were dissected, fixed overnight in 4% formaldehyde in PBS at 4°C, dehydrated and embedded in paraffin. 10 μm sections were rehydrated, treated 15 min with 6% H₂O₂, 10 min with 20 μg/ml proteinase K, post-fixed with 4% formaldehyde for 15 min, treated with acetic anhydride, equilibrated with 2× SSC and hybridized overnight at 63°C with digoxygenin labeled RNA probes for Axin2, Kcne3 and Sfrp5. Sections were washed and incubated overnight with sheep anti-digoxygenin antibody 1:3000 (Roche). The signal was revealed using NBT/BCIP (Roche). Images were acquired with Leica DM2500 microscope. For antibody staining, small intestines were dissected, fixed for 20 min in 1% formaldehyde in PBS at 4°C, incubated overnight in 30% sucrose, embedded in OCT and kept at −80°C. Immunohistochemical analyses were performed on 10 μm cryosections using 1:1000 anti-EpCAM antibody (rat, eBiosciences), anti-H2A.Z 1:1000 (Active Motif, #39113) and H3K27Ac 1:1000 (Abcam 4729), followed by 1:300 Alexa566-conjugated secondary antibody (Invitrogen), Images were acquired with Leica AF7000 microscope.