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c-Fos Monoclonal Antibody

  • Cat.No.:E-AB-70204

  • 宿主: Mouse
  • 反应性: H,M,R
  • 应用: WB,IHC,IF

订购:E-AB-70204

规格:
  • 60μL
  • 120μL
价格: ¥1420
数量:

Test Application

  • Verified Samples

    Reactivity Application
    Human WB
    (HepG2,Jurkat,colon cancer,)

    Western Blot analysis of various samples using c-Fos Monoclonal Antibody at dilution of 1:1000.

    Western Blot analysis of various samples using c-Fos Monoclonal Antibody at dilution of 1:1000.

    Western Blot analysis of various samples using c-Fos Monoclonal Antibody at dilution of 1:1000.

    IHC
    (gallbladder,)

    Immunohistochemistry analysis of paraffin-embedded Human gallbladder using c-Fos Monoclonal Antibody at dilution of 1:1000.

    Rat IF
    (brain,)

    Immunofluorescence analysis of paraffin-embedded Rat brain using c-Fos Monoclonal Antibody at dilution of 1:400.

    Mouse WB
    (lung,)

    Western Blot analysis of various samples using c-Fos Monoclonal Antibody at dilution of 1:1000.

    IHC
    (brain,)

    Immunohistochemistry analysis of paraffin-embedded Mouse brain using c-Fos Monoclonal Antibody at dilution of 1:1000.

  • Dilution

    WB 1:500-1:2000, IHC 1:500-1:2000, IF 1:200-1:1000

蛋白样本的制备

1.样本的处理

1)组织样本的处理:
a) 取待测组织样本,用预冷的PBS(0.01 M, pH=7.4)(Cat# E-BC-R187) 充分洗涤,洗去组织表面血液及内部杂物。
b) 称重剪碎,加入适量比例的RIPA裂解液混合物 (Cat# E-BC-R327)(1mL的RIPA裂解液中加入10μL PMSF(Cat# E-BC-R287)和10μL原矾酸钠(Cat# E-BC-R250)进行匀浆裂解。建议按组织重量:RIPA体积=3:10的比例匀浆,例如0.3g的组织样本对应1mL的RIPA裂解液,具体体积可根据实验需要适当调整。
c) 在35~40%的功率下,超声处理样本1min(冰浴条件下进行),每次2s,间隔2s,确保细胞充分裂解,降低样本粘度。
d) 4℃下12,000rpm离心10min。
e) 取上清,待测定蛋白浓度。

2)细胞样本处理:
a) 收集待测细胞样本,用预冷的PBS(0.01 M, pH=7.4)充分洗涤,洗去培养基等成分(一般建议洗涤3次)。
b) 加入适量比例的RIPA裂解液混合物(1mL RIPA裂解液中加入10μL PMSF和10μL原矾酸钠)进行冰上裂解。建议6孔板中每孔加入0.1mL RIPA裂解液(细胞中的蛋白含量可能会不同,可适当调整加入裂解液的体积)。
c) 在35~40%的功率下,超声处理样本1min(冰浴条件下进行),每次2s,间隔2s,确保细胞充分裂解,降低样本粘度。
d) 4℃下12,000rpm离心10min。
e) 取上清,待测定蛋白浓度。

2.BCA法测定蛋白浓度(参见BCA蛋白浓度测定试剂盒(Cat# E-BC-K318)说明书)。

3. 用PBS调整蛋白浓度,按照蛋白样本:5×SDS上样缓冲液=4:1的比例加入5×SDS上样缓冲液(Cat# E-BC-R288),沸水煮10min。12,000rpm离心2min,收取上清。变性后的蛋白即可进行后续的Western Blot实验,或保存于 -20℃或-80℃。

电泳

1.根据靶蛋白的分子量的大小,配制 0% 的分离胶。每个泳道加入待测样本,并预留一个泳道加入5μL的预染蛋白Marker (Cat# E-BC-R273),以验证转膜程度及目的分子量大小。加入电泳缓冲液 ( Cat# E-BC-R331),开始电泳。

2.80V恒压电泳,待溴酚蓝指示剂移动至浓缩胶与分离胶交界处成线状,改为恒压120v,跑完全程。

转膜

1.根据靶蛋白的分子量选择 μm 孔径的PVDF膜(Cat# E-BC-R266)。PVDF膜先在甲醇中浸泡1min使其活化,然后将PVDF膜浸泡于转膜缓冲液(Cat# E-BC-R333)中。同时将滤纸和纤维垫也浸泡在转膜缓冲液中待用。

2.遵照转膜仪生产商的说明进行湿转、半干转或干式转印。

免疫印迹

1.用含5% 脱脂奶粉(Cat# E-BC-R337)的TBST(Cat# E-BC-R335)(封闭液)浸泡PVDF膜,室温摇床封闭

2.用含5% 脱脂奶粉的TBST溶液按照说明书推荐的稀释比选取稀释IMP3抗体,使PVDF膜浸泡于一抗孵育液中,4℃孵育过夜,并不断轻轻晃动。

3.TBST充分洗涤PVDF膜 .

4.用含2%脱脂奶粉的TBST溶液(配比见附录)按照说明书推荐的稀释比选取稀释Goat Anti-Rabbit IgG(H+L)(peroxidase/HRP conjugated)(Cat# E-AB-1003),室温摇床孵育1 h。

5.TBST充分洗涤PVDF膜 .

显色曝光

1.PVDF膜从TBST洗液中取出后用滤纸吸干水分。

2.将ECL化学发光检测试剂盒(Cat# E-BC-R347)中的A液与B液按1:1比例混匀,均匀滴加于PVDF膜上,排出气泡,即可曝光。

3.调节对比度,多次曝光获取最佳图片效果。

附录

Product Details

Clonality Monoclonal
Isotype IgG
Concentration 0.97 mg/mL
Storage Store at -20℃. Avoid freeze / thaw cycles.
Buffer PBS with 0.02% sodium azide,100 μg/ml BSA and 50% glycerol.
Purification Method Affinity purification
Research Areas Cancer, Epigenetics and Nuclear Signaling, Neuroscience
Clone No. 4B4F11
Conjugation Unconjugated

Immunogen Details

Immunogen KLH conjugated Synthetic peptide corresponding to Mouse c-FOS
Abbre c-Fos
Swissprot P01100,P01101,P12841
Calculated MW 62kDa
Observed MW 62kDa

Background

The Fos gene family consists of 4 members: FOS, FOSB, FOSL1, and FOSL2. These genes encode leucine zipper proteins that can dimerize with proteins of the JUN family, thereby forming the transcription factor complex AP-1. As such, the FOS proteins have been implicated as regulators of cell proliferation, differentiation, and transformation. In some cases, expression of the FOS gene has also been associated with apoptotic cell death.

Citations

  1. Life sciences(2019)IF: 3.647
    miR-A-181a over-expression in mesenchymal stem cell-derived exosomes influenced inflammatory response after myocardial ischemia-reperfusion injury

    DOI: 10.1016/j.lfs.2019.116632

    PMID: 31278944

    物种: Human,Mouse 样本类型: Cell lysate,Tissue homogenate

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