APC Rabbit Recombinant mAb

Filter:

规格 价格 库存 购买数量
20ul RMB 447.8 现货
100ul RMB 1500.11 现货
更大包装 有超大折扣 点击询价
 

400-668-6834

info@selleck.cn

 

使用信息

抗体应用 WB, IF,ELISA
稀释比例
WB IF
1:500 1:50
反应性 Human Rat
MW (kDa) 160kDa
抗体类型 Rabbit
浓度 1mg/ml
储存液配方 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide.
储存条件
(自收到货起)
Store at –20°C.

Datasheet & SDS

生物描述

特异性 APC Rabbit Recombinant mAb detects endogenous level of total APC.
背景 The adenomatous polyposis coli (APC) protein is a multi-domain protein that contains binding sites for numerous proteins, including microtubules, the Wnt/Wg pathway components β-catenin and axin, the cytoskeletal regulators EB1 and IQGAP1, and the Rac guanine-nucleotide-exchange factor (GEF) Asef1. It is atumor suppressor involved in several cellular processes and signal transduction pathways such as the canonical Wnt signal transcription, cell adhesion, cell migration and chromosomal segregation. APC plays a central role suppressing the canonical Wnt signalling pathway that controls cell proliferation and differentiation in various organs. APC inhibits β-catenin/TCF-dependent transcription through multiple mechanisms. It could provide a scaffold for a destruction complex together with GSK3β and axin that promotes phosphorylation and consequent ubiquitin-dependent degradation of β-catenin. Second, APC promotes export of β-catenin from the nucleus, which reduces the amount of nuclear β-catenin/TCF. Third, APC binds to β-catenin, blocking the interaction with TCF. Finally, recent studies show that APC can inhibit β-catenin/TCF-dependent transcription through a direct interaction with a repressor complex.

实验方法

WB

Western Blotting

Sample preparation

1. Aspirate media from cultures and Wash the cells with 1X PBS.
2. Lyse cells by adding 1X SDS sample buffer and transfer the extract to a microcentrifuge tube. Keep onice.
3. Sonicate for 10–15 sec to complete cell lysis and shear DNA.
4. Heat a 20 µl sample to 95–100°C for 5 min, then cool on ice.
5. Centrifuge for 5 min (with Microcentrifuge).
6. Load appropriate volumes of samples onto SDS-PAGE gel (loading quantity of protein sample depends on the concentration of extracted proteins).
NOTE: At the same time, please load the pre-stained molecular weight markers to determine molecular weights and verify electrotransfer.
7. Electrotransfer to nitrocellulose/PVDF membrane.

Membrane Blocking and Antibody Incubations

a. Blocking

1. (Optional) After transfer, wash the transferred membrane with TBS for 5 min at room temperature.
2. Incubate the membrane in the blocking buffer for 1 hr at room temperature.
3. Wash three times for 5 min each with TBST.

b. Antibodies Incubation

1. Incubate membrane and primary antibody (at the appropriate dilution and diluent recommended) in a primary antibody dilution buffer with gentle agitation overnight at 4°C.
2. Wash three times for 5 min each with TBST.
3. Incubate membrane with an appropriate second antibodydissolved in the blocking buffer with gentle agitation for 1 hr at room temperature.
4. Wash three times for 5 min each with TBST.
5. Proceed with detection.

Detection of Proteins

1. After antibodies incubation, Wash membrane three times for 5 minutes in TBST.
2. PrepareECL Reagent (or other chromogenic agents/substrate according to your second antibody). Mix well.
3. Incubate substrate with membrane for 1 minute, remove excess solution (membrane remains wet), wrap in plastic and expose to X-ray film.

IF

Immunofluorescence (Immunocytochemistry)

Specimen Preparation (forcultured cell lines, IF-IC)

1. Aspirate liquid, then cover cells to a depth of 2–3 mm with 4% formaldehyde diluted in 1X PBS.
NOTE: Formaldehyde is toxic, use only in a fume hood.
2. Fix cells for 15 min at room temperature.
3. Aspirate fixative, rinse three times in 1X PBS for 5 min each.
4. Proceed with Immunostaining.

Immunostaining

1. Add theblocking buffer and incubate for 60 min at RT.
2. Prepare primary antibody diluent in antibody dilution buffer as recommended .
3. Aspirate blocking solution, apply diluted primary antibody.
4. Incubate overnight at 4°C.
5. Rinse three times in 1X PBS for 5 min each.
6. Incubate specimens in fluorochrome-conjugated secondary antibody diluted in antibody dilution buffer for 1–2 hr at room temperature in the dark.
7. Rinse three times in 1X PBS for 5 min each.
8. Mount slides usingmounting medium with DAPI and cover with coverslips.
9. For best results, allow mountant to cure overnight at room temperature. For long-term storage, store slides flat at 4°C protected from light.

技术支持

在订购、运输、储存和使用我们的产品的任何阶段,您遇到的任何问题,均可以通过拨打我们的热线电话400-668-6834,或者技术支持邮箱tech@selleck.cn,直接联系到我们。我们会在24小时内尽快联系您。

* 必填项

请输入您的姓名
请输入您的邮箱地址 请输入一个有效的邮箱地址
请写点东西给我们
在线咨询
联系我们