WB, Western blot; IP, Immunoprecipitation; IF, Immunofluorescence; IHC, Immunohistochemistry; ICC, Immunocytochemistry; FC, Flow Cytometry; ChIP, Chromatin Immunoprecipitation Assay; ChIP-seq, ChIP-sequencing.

H, Human; M, Mouse; R, Rat; Ar, Arabidopsis; B, Bovine; C, Chicken; Cerastium, Ce; Cw, Cow; Dg, Dog; Gp, Guinea pig; Hm, Hamster; Hr, Horse; Mk, Monkey; Ph, Pharbitis; Pg, Pig; Rb, Rabbit; Rc, Rice; S, Sheep; Z, Zebrafish; All, all species expected.

配套提供了Western一抗稀释液，可以用于Western检测或其它适当用途时的一抗稀释。

建议抗体使用时的稀释比例如下(实际使用时需根据抗原水平的高低作适当调整)：

ChIP-seq 1:1,000-1:2,000 1:800-1:2,000 1:800-1:2,000 1:200-1:1,000

抗体详细信息如下:

Background Epitope tags are useful for the labeling and detection of proteins using immunoblotting, immunoprecipitation, and immunostaining techniques. Because of their small size, they are unlikely to affect the tagged protein's biochemical properties. A variety of plasmids contain DNA that encodes an amino-terminal tag consisting of six histidine (6x His) residues followed by an extended multiple cloning site. The 6x His tag on the expressed recombinant proteins allows for efficient coupling to Ni2+ affinity resins and purification by single step chromatography. As is the case with other protein tag systems, this polyhistidine tag can often be cleaved at sites recognized by proteases such as thrombin and enterokinases to isolate the protein of interest. 保存条件：

His Tag Rabbit Monoclonal Antibody -20℃保存，需避免反复冻融。Western一抗稀释液-20℃或4℃保存，一年有效，Western一抗稀释液优先推荐4℃保存，长期不使用可以考虑-20℃保存，但冻融可能会导致出现轻微的浑浊和少量不溶物。

注意事项：

如果本抗体用于Western blot (WB)、免疫荧光(IF)、免疫细胞化学(ICC)等实验，请注意回收使用过的稀释抗体。回收的抗体通常至少可以重复使用5-10次。稀释后的抗体，包括已经使用过的稀释抗体，请4℃保存。

回收后重复使用的抗体，使用方法同新鲜稀释的抗体。如果在重复使用过程中发现抗体出现轻微混浊现象，可以10,000×g离心1-3分钟，取上清用于后续检测。如果回收的抗体出现明显的絮状物或长霉长菌等情况，则可以考虑废弃该抗体。

提供的Western一抗稀释液也可以用于免疫荧光(IF)、免疫组化(IHC)、免疫细胞化学(ICC)等适当用途。如果希望获得最佳的检测效果，请考虑使用上述检测专用的一抗稀释液。

本产品仅限于专业人员的科学研究用，不得用于临床诊断或治疗，不得用于食品或药品，不得存放于普通住宅内。

为了您的安全和健康，请穿实验服并戴一次性手套操作。

1.Western检测：
a.按照推荐的稀释比例用碧云天提供的Western一抗稀释液稀释抗体。
b.把经过封闭的蛋白膜与稀释好的一抗4℃缓慢摇动过夜或室温缓慢摇动2小时，确保稀释的抗体至少能在摇动的瞬间覆盖蛋白膜。
c.回收稀释的一抗，4℃保存以备下次继续使用。
d.按照Western的实验步骤进行后续的洗涤、二抗孵育、洗涤和检测等操作。具体操作可以参考如下网页：
http://www.beyotime.com/support/western.htm
2. 免疫染色：
可以使用碧云天生产的 免疫染色一抗稀释液(P0103) 稀释抗体，使用后注意回收稀释好的一抗，具体操作可以参考如下网页： http://www.beyotime.com/support/immunol-staining.htm
3. 其它实验操作请自行参考适当的protocol进行。
4. 代表性图片。
All lanes: Anti-His tag antibody at 1:2,000 dilution Observed MW: Depend on the fusion protein with His tag Lane 1: fusion protein with N-terminal 6X His tag Lane 2: fusion protein with C-terminal 6X His tag Lane 3: Multi-tag fusion protein containing 6X His tag Lane 4: 293 cells lysate-transfected with N-terminal 6X His tagged gene Lane 5: 293 cells lysate-transfected with C-terminal 6X His tagged gene Lane 6: fusion protein with N-terminal 6X His tag Lane 7: fusion protein with C-terminal 6X His tag Lane 8: untransfected 293 cell lysate Lysate at 20 μg per lane 2nd Ab: GAR HRP(H+L) 1:5,000 Exposure: 60s His tag was immunoprecipitated from 0.05mg of 293 whole cells lysate transfected with N-terminal 6X His tagged gene with His-tag Rabbit Monoclonal Antibody at 1:50 dilution. 2nd Ab: GAR HRP for IP 1:500 Lane 1: His-tag Rabbit Monoclonal Antibody IP in 293 whole cell lysate transfected with N-terminal 6X His tagged gene Lane 2: PBS instead of His-tag Rabbit Monoclonal Antibody in 293 whole cell lysate transfected with N-terminal 6X His tagged gene Lane 3: 293 whole cell lysate transfected with N-terminal 6X His tagged gene, 10μg (input) Exposure: 20s His-tag Rabbit Monoclonal Antibody staining 293 cells trasnfected with C-terminal 6X His tagged gene by IF/ICC . Cells were fixed with paraformaldehyde, permeabilized with 0.1% Triton X-100 and blocked with 10% goat serum for half an hour at room temperature. Samples were incubated with primary antibody (1:800) at 4°C. An Alexa Fluor® 488-conjugated Goat Anti-Rabbit IgG polyclonal was used as the secondary antibody (1:500). DAPI (blue) was used as the nuclear counter stain. Control: PBS and secondary antibody, An Alexa Fluor® 488-conjugated Goat Anti-Rabbit IgG (1:500). Overlay histogram showing 293 cells transfected with N-terminal (Blue) and C-terminal (Red) 6X His tagged gene stained with His-tag Rabbit Monoclonal Antibody. The cells were then incubated in the antibody (His-tag Rabbit Monoclonal Antibody, 1:1,000 dilution) in 1x PBS/1% BSA for 30 min at at room temperature. The secondary antibody used was a Goat Anti-Rabbit Alexa Fluor® 488 (IgG H+L) at 1:2,000 dilution for 20 min at room temperature. Unlabelled sample (Black) was used as a control.