After washing in the same buffer, the cells were permeabilized with a 0.2% solution of Triton X-100 detergent (Sigma) in PBS for 1 minute at room temperature.
After washing with 0.01 M phosphate buffer, the cells were permeabilized in PBS containing 0.5% Triton X-100 for 10 min at room temperature and then washed three times with PBS.
After two washes with Perm/Wash buffer (BD Biosciences), the cells were permeabilized with Cytoperm permeabilization buffer (BD Biosciences) for 10 min at 4°C, washed with Perm/Wash buffer (BD Biosciences), and subjected to DNase (300 μg/ml in PBS) digest for 1 h at 37°C.
Hippocampal neurons were fixed with 4% paraformaldehyde in phosphate buffer (PB; pH 7.4) for 15 min. After washing with 1x phosphate buffer saline (PBS), cells were permeabilized with 0.05% Triton X-100 in 1xPBS for 15 min, and incubated with blocking buffer (3% BSA, 0.02% Triton X-100 in 1xPBS) for 15 min before being probed with primary antibodies.
After washing 3 times in phosphate buffered saline (PBS) cells were permeabilized with 0.2% triton X-100 for 5 minutes at room temperature, followed by 3 further PBS washes.
Thereafter, cells were permeabilized using permeabilization buffer (eBioscience) and stained with PE-IL-10 (BD Pharmingen), or appropriate isotype control.
To determine GATA3 serine phosphorylation, cells were permeabilized with permeabilization buffer (PB) (PBS 10% FCS 0.5% saponine) and labeled with anti-pSer308-GATA3 anti-pSer308-GATA3 anti-pSer308-GATA3per 1 x 10 cells) followed by incubation with anti-rabbit IgG-Alexa 488.
Cells were permeabilized with a permeabilization buffer (eBioscience) before staining with ALDH1, collagen type II, nestin, βIII-tubulin and GFAP.
Briefly, the slides were rinsed with wash buffer (PBS with 0.5%Tween-20Tween-20the cells were permeabilized with 0.25% Triton for 10 min.
For chromatin fractionation experiments, cells were permeabilized in RSB buffer containing 10 mM Hepes-NaOH (pH 7.5), 1.5 mM MgCl2, 0.5 mM EDTA, 10 mM KCl, 0.5%NP400, phosphatase, and protease inhibitors.
