Phospho-PAKα (T212) Polyclonal Antibody

Phospho-PAKα (T212) Polyclonal Antibody发货快，折扣优，省时省力！
宿主兔
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克隆类型多克隆
应用范围
Immunohistochemistry (Frozen Sections) (IHC (fro)), Immunohistochemistry (Paraffin-embedded Sections) (IHC (p)), Immunohistochemistry (IHC), Western Blotting (WB)

Phospho-PAKα (T212) Polyclonal Antibody实验流程    
Harvest Brain from 13 weeks old mice in 1x Dulbecco’s phosphate buffered saline (DPBS) (Gibco Life Technologies, 14200-067).
Fix mouse brain in antigen fix (Diapath, P0014) for 2h at RT.
Wash tissue in 0.1M pH7.4 phosphate buffer at for 1h at 4°C.
Dehydrate tissue in 30% sucrose solution ON at 4°C.
Snap freeze tissue in Tissue Freezing Medium (ElectroMicroscopy Science, 72592-C) at -80°C.
Cut blocks into 20μm sections using a cryostat (Leica, CM3050 S).
Transfer sections to a slide.
Create a hydrophobic barrier on the slide around sections with Dako pen (Dako, S2002, lot 00081640).
Place slide in a humidified chamber and rehydrate sections in 0.1M TrisHCl pH7.4 for 10min at RT.
Gently remove buffer by tapping slide.
Permeabilize tissue in 0.1M TrisHCl pH7.4 containing 2% Triton X-100 (Sigma, batch 015K0039) and 0.5% BSA, for 20min at RT.
Incubate sections with primary AIF1 (c-term) (antibodies-online, ABIN2857032, Lot 39476) diluted 1:500 in 0.1M TrisHCl pH7.4 containing 2% Triton X-100 (Sigma, batch 015K0039) and 0.5% BSA for 5h at RT. Incubate the negative contrls sections with diluted rabbit serum.
Wash slides 1x 5min in 0.1M TrisHCl pH7.4.
Incubate sections with secondary donkey Fab’2 anti-rabbit A647 conjugated antibody (Jackson Immunoresearch, 711-606-152, lot 128806) diluted 1:500 in 0.1M TrisHCl pH7.4 containing 2% Triton X-100 and 0.5% BSA ON at RT.
Wash slides 1x 5min in 0.1M TrisHCl pH7.4.
Add approximately 10μl of Slowfade? Gold antifade reagent (Life technologies, 536937, lot 1226836) for each section and mount cover slip.
Image acquisition on a LSM 880 (Zeiss), 20x magnification, 1000 resolution.