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Arf6活性检测试剂盒
Arf6活性检测试剂盒图片
型号: Arf6
货号: 82401
品牌: NewEast Biosciences
参考价格: 6800元/盒
包装: 30 Assays
交货期: 现货


包装与价格:
订货号 纯度规格 包装 价格(元)
82401 30 Assays 6800
半岛bd体育手机客户端 名称
Arf6 Pull-Down Activation Assay Kit
半岛bd体育手机客户端 介绍

Arf6 Pull-Down Activation Assay Kit

Cat. # 82401

Introduction

A. Background
ARF6 (ADP-ribosylation factor 6) is a member of the ARF super-family. ARF genes encode small GTPases that increase the ADP-ribosyltransferase activity of cholera toxin and are critical for vesicular trafficking as activators of phospholipase D. Arf6 regulates membrane trafficking and functions as a regulatory molecule of phagocytosis.
Currently there is no direct assay to measure the activation of Arf6 GTPase.
B. Assay Principle
The Arf6 Activation Assay Kit uses configuration-specific anti-Arf6-GTP Mouse monoclonal antibody to measure Arf6-GTP levels in cell extracts or in vitro GTPγS loading Arf6 activation assays. Anti-Arf6-GTP mouse monoclonal antibody is first incubated with cell lysates containing Arf6-GTP. Next, the GTP-bound Arf6 is pulled down by protein A/G agarose. Finally, the precipitated Arf6-GTP is detected through immunoblot analysis using Anti-Arf6 Rabbit Polyclonal Antibody.
The anti-Arf6-GTP monoclonal antibody can also be used to monitor the activation of Arf6 in cells and in tissues by immunohistochemistry.
C. Kit Components
1. Anti-Arf6-GTP Mouse Monoclonal Antibody (Cat. # 26918): One vial – 35 μL (1 mg/ml) in PBS, pH 7.4, containing 50% glycerol. This antibody specifically recognizes Arf6-GTP from all vertebrates.
2. Protein A/G Agarose (Cat. # 30301): One vial – 600 μL of 50% slurry.
3. 5X Assay/Lysis Buffer (Cat. # 30302): One bottle – 30 mL of 250 mM Tris-HCl, pH 8, 750 mM NaCl, 50 mM MgCl2, 5 mM EDTA, 5% Triton X-100.
4. Anti-Arf6 Rabbit Polyclonal Antibody (Cat. # 21032): One vial – 50 μL (1 mg/mL) in PBS, pH 7.4, contained 50% glycerol.
5. 100X GTPγS (Cat. # 30303): One vial – 50 μl at 10 mM, use 5 μL of GTPγS for GTP-labeling of 0.5 mL of cell lysate.
6. 100X GDP (Cat. # 30304): One vial – 50 μl at 100 mM, use 5 μL of GDP for GDP-labeling of 0.5 mL of cell lysate.
7. HRP-Goat Anti-Rabbit IgG (Cat. #29002): 50 μL (0.4 mg/mL) in PBS, pH 7.4, contained 50% glycerol.
D. Materials Needed but Not Supplied
1. Stimulated and non-stimulated cell lysates
2. Protease inhibitors
3. 4 °C tube rocker or shaker
4. 0.5 M EDTA at pH 8.0
5. 1.0 M MgCl2
6. 2X reducing SDS-PAGE sample buffer
7. Electrophoresis and immunoblotting systems
8. Immunoblotting wash buffer such as TBST (10 mM Tris-HCl, pH 7.4, 0.15 M NaCl, 0.05% Tween-20)
9. Immunoblotting blocking buffer (TBST containing 5% Non-fat Dry Milk or 3% BSA)
10. ECL Detection Reagents


E. Example Results
The following figure demonstrates example results seen with the Arf6 Activation Assay Kit. For reference only.

Arf6 Activation Assay. Purified Arf6 proteins were immunoprecipitated after treated with GDP (lane 1) or GTPγS (lane 2). Immunoprecipitation was done with the anti-Arf6-GTP monoclonal antibody (Cat. No. 26918). Immunoblot was with an anti-Arf6 rabbit polyclonal antibody (Cat. No. 21032).

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