Cat.No.:E-AB-70075
Reactivity | Application |
Human | IHC (brain,) ![]() Immunohistochemistry analysis of paraffin-embedded Human brain using Tau Polyclonal Antibody at dilution of 1:100. |
Rat |
WB (brain,) ![]() Western Blot analysis of various samples using Tau Polyclonal Antibody at dilution of 1:1000. (brain,) ![]() Immunohistochemistry analysis of paraffin-embedded Rat brain using Tau Polyclonal Antibody at dilution of 1:100. |
Mouse |
WB (brain,) ![]() Western Blot analysis of various samples using Tau Polyclonal Antibody at dilution of 1:1000. |
WB 1:500-1:2000, IHC 1:100-1:500
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℃。
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。
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。
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 .
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。
Clonality | Polyclonal |
Isotype | IgG |
Concentration | 150 μg/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, Neuroscience, Signal Transduction |
Conjugation | Unconjugated |
Immunogen | KLH conjugated Synthetic peptide corresponding to Mouse Tau |
Abbre | Tau |
Synonyms | MAPT, DDPAC, FTDP-17, MAPTL, MSTD, MTBT1, MTBT2, PPND, PPP1R103 |
Swissprot | P10636,P10637,P19332 |
Calculated MW | 40-70kDa |
Observed MW | 40-70kDa |
Cellular Localization | Cytoplasm>cytosol. Cell membrane. Cytoplasm>cytoskeleton. Cell projection>axon. Mostly found in the axons of neurons, in the cytosol and in association with plasma membrane components. |
Tissue Specificity | Expressed in neurons. Isoform PNS-tau is expressed in the peripheral nervous system while the others are expressed in the central nervous system.Highest expression level in parietal lobe. |
Application | Products |
WB | Western Blot Detection Kit(E-IR-R304) |
IHC | 2-step plus Poly-HRP Anti Mouse/Rabbit IgG Detection System (with DAB solution)(E-IR-R217) |
This gene encodes the microtubule-associated protein tau (MAPT) whose transcript undergoes complex, regulated alternative splicing, giving rise to several mRNA species. MAPT transcripts are differentially expressed in the nervous system, depending on stage of neuronal maturation and neuron type. MAPT gene mutations have been associated with several neurodegenerative disorders such as Alzheimer's disease, Pick's disease, frontotemporal dementia, cortico-basal degeneration and progressive supranuclear palsy. |