Menzel glass

We prepare Trichloro (1H,1H,2H,2H-perfluorooctyl)silane (PFOTS) and trichloro(octyl)silane (OTS) on 1 mm thick Menzel glass (Thermo Fisher Scientific Ltd) by a chemical vapor deposition process.

Sections were washed three times in KPBS for 10 min and imbedded in a solution containing 300 g/L egg-albumin (Sigma-Aldrich) and 30 g/L gelatin (Sigma-Aldrich), frozen at −80 °C and sub-sliced on a cryotome at −20 °C (Cellab Nordia AB) to 16 μm thick slices. Slices were mounted on positively charged glasses (+Menzel glass, Thermo Scientific) and stored at −20 °C. These slices were then preincubated in 10% Donkey serum, 0.25% Triton X-100 in KPBS for 1 h. After adding rabbit anti-NPY antibody (1:500, #N9528, Sigma-Aldrich, DK) in 5% Donkey serum, 0.25% Triton X-100 in KPBS, sections were incubated overnight at 4 °C. Sections were washed 3 × 10 min in tKPBS, incubated for 2 h at RT with Cy3-conjugated Donkey anti-rabbit antibody (1:200, Jackson Immunoresearch USA) in 1% Donkey serum, 0.25% Triton X-100 in KPBS and washed 1 × 10 min in tKPBS and 2 × 10 min in KPBS. The slides were mounted with DABCO (Sigma-Aldrich) and digitized images obtained using Olympus BX61 microscope and CellSens software.