HDAC1 Antibody (10E2): sc-81598

Protocol tips

Upstream tips	Protocol tips	Downstream tips
	-Mouse (1:500) -Use clean forceps to gently handle the blot from the corner without creasing the membrane. Do not write on the blot with pen or marker, as the ink can fluoresce and cause background.

Protocol tips
-Mouse (1:500) -Use clean forceps to gently handle the blot from the corner without creasing the membrane. Do not write on the blot with pen or marker, as the ink can fluoresce and cause background.

Publication protocol

Proteins were separated by SDS-PAGE and transferred on Immobilon-P transfer membrane (IPVH00010, Merck Millipore). After blocking with TBST (25 mM Tris pH 8.0, 150 mM NaCl, and 0.1% Tween 20) containing 6.25% (w/v) nonfat dry milk for 90 min, the membrane was reacted with a primary antibody for 1 h. The following primary antibodies were used in this study: anti-myc (2 μg/ml), anti-FcεRIβ (1:1000, a gift from Dr. Reuben P. Siraganian, National Institutes of Health, Bethesda, MD), anti-FcεRIγ (1:300, 06-727), anti-phosphotyrosine (pY) (1:1000, clone 4G10, 05-321), anti-GAPDH (1:1000, MAB374, Merck Millipore), anti-phosphoERK (Thr202/Tyr204) (9101S), anti-ERK (9102S), anti-phosphoSyk (Tyr525/526) (2710P), anti-Syk (2712P), anti-phosphoPLCγ2 (Tyr1217) (3871P) (1:500, Cell Signaling Technology, Danvers, MA), anti-PLCγ2 (Q-20), anti-NFATc1 (7A6), anti-NFATc2 (4G6-G5), and anti-histone deacetylase 1 (HDAC1) (10E2) (1:500, Santa Cruz Biotechnology, Santa Cruz, CA). After extensive washing, the membranes were reacted with horseradish peroxidase-conjugated anti-mouse (115-035-146) or anti-rabbit (111-035-003) secondary antibodies (1:10000, Jackson ImmunoResearch, West Grove, PA) for 30 min. Proteins were visualized using enhanced chemiluminesence reagent (Western Lightning Plus-ECL, Perkin Elmer Life Sciences, Waltham, MA). The antibodies bound to membrane were stripped and reprobed with the other antibodies as described1

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