Goat anti rabbit a488 secondary antibody

Fluorescence microscopy was performed as previously described^{81 (link)}. Adult male mice (six ePet-Rgs2^lo mice, five ePet-Rgs2^hi mice) were perfused with 4% paraformaldehyde in phosphate-buffered saline (PBS), pH 7.4, and dissected brains post fixed in the same solution for 1 h. Brains were cryoprotected in 30% sucrose (wt vol⁻¹), 1× PBS overnight at 4 °C. Thirty micrometers of coronal cryostat sections were collected, blocked in 5% fetal bovine serum (FBS) (fetal bovine serum) in PBST (PBS with 0.1% Triton X-100) for 1 h at room temperature and incubated with a primary antibody against GFP (1:1000 dilution, Frontier Institute) and conjugated with goat-anti-rabbit-A488 secondary antibody (1:1000; Molecular Probes). Images of brain slices were analyzed with a Leica TCS SP5II confocal microscope using a ×20/0.3 NA objective. The intensity of the GFP fluorescence in serotonergic neurons was quantified using NIH Image J.

MDCK and A549 cells were washed, fixed with formaldehyde, permeabilized with 0.5% triton and labeled with a rabbit anti-ZO1 primary antibody (Zymed) and goat anti-rabbit A488 secondary antibody (Molecular Probes). Nuclei were labeled with Hoechst 33258 (10 μg/mL, Sigma-Aldrich). Preparations were observed with an Olympus inverted microscope with a 20x air objective.