重组Anti-Chk2抗体[EPR4325] - BSA and Azide free (ab227998)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR4325] to Chk2 - BSA and Azide free
- Suitable for: IP, IHC-P, WB, Flow Cyt (Intra), ICC/IF
- Knockout validated
- Reacts with: Human
Related conjugates and formulations
概述
-
产品名称
Anti-Chk2抗体[EPR4325] - BSA and Azide free
参阅全部 Chk2 一抗 -
描述
兔单克隆抗体[EPR4325] to Chk2 - BSA and Azide free -
宿主
Rabbit -
经测试应用
适用于: IP, IHC-P, WB, Flow Cyt (Intra), ICC/IFmore details -
种属反应性
与反应: Human -
免疫原
Recombinant fragment. This information is proprietary to Abcam and/or its suppliers.
-
阳性对照
- WB: HeLa (untreated and treated with gamma irradiation), HEK-293, MDA-MB-231, HT-29, and 293T cell lysates. IHC-P: Human colon and spleen tissues. ICC/IF: Wild-type HAP1 cells. IP: HeLa whole cell lysate.
-
常规说明
ab227998 is the carrier-free version of ab109413.
Our carrier-free antibodies are typically supplied in a PBS-only formulation, purified and free of BSA, sodium azide and glycerol. The carrier-free buffer and high concentration allow for increased conjugation efficiency.
This conjugation-ready format is designed for use with fluorochromes, metal isotopes, oligonucleotides, and enzymes, which makes them ideal for antibody labelling, functional and cell-based assays, flow-based assays (e.g. mass cytometry) and Multiplex Imaging applications.
Use our conjugation kits for antibody conjugates that are ready-to-use in as little as 20 minutes with <1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.
This product is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm, without the need for antibody preparation. Maxpar® is a trademark of Fluidigm Canada Inc.
Mouse, Rat: We have preliminary internal testing data to indicate this antibody may not react with these species. Please contact us for more information.
性能
-
形式
Liquid -
存放说明
Shipped at 4°C. Store at +4°C. Do Not Freeze. -
存储溶液
pH: 7.20
Constituent: PBS -
无载体
是 -
Concentration information loading...
-
纯度
Protein A purified -
克隆
单克隆 -
克隆编号
EPR4325 -
同种型
IgG -
研究领域
相关产品
-
Alternative Versions
-
Compatible Secondaries
-
Conjugation kits
-
Isotype control
-
KO cell lines
-
KO cell lysates
-
Positive Controls
应用
The Abpromise guarantee
Abpromise™承诺保证使用ab227998于以下的经测试应用
“应用说明”部分 下显示的仅为推荐的起始稀释度;实际最佳的稀释度/浓度应由使用者检定。
应用 | Ab评论 | 说明 |
---|---|---|
IP |
Use at an assay dependent concentration.
|
|
IHC-P |
Use at an assay dependent concentration. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
antigen retrieval is recommended. |
|
WB |
Use at an assay dependent concentration. Detects a band of approximately 62 kDa (predicted molecular weight: 61 kDa).
|
|
Flow Cyt (Intra) |
Use at an assay dependent concentration.
ab199376 - Rabbit monoclonal IgG (Low endotoxin, Azide free), is suitable for use as an isotype control with this antibody. |
|
ICC/IF |
Use at an assay dependent concentration.
|
说明 |
---|
IP
Use at an assay dependent concentration. |
IHC-P
Use at an assay dependent concentration. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol. antigen retrieval is recommended. |
WB
Use at an assay dependent concentration. Detects a band of approximately 62 kDa (predicted molecular weight: 61 kDa). |
Flow Cyt (Intra)
Use at an assay dependent concentration. ab199376 - Rabbit monoclonal IgG (Low endotoxin, Azide free), is suitable for use as an isotype control with this antibody. |
ICC/IF
Use at an assay dependent concentration. |
靶标
-
功能
Regulates cell cycle checkpoints and apoptosis in response to DNA damage, particularly to DNA double-strand breaks. Inhibits CDC25C phosphatase by phosphorylation on 'Ser-216', preventing the entry into mitosis. May also play a role in meiosis. Regulates the TP53 tumor suppressor through phosphorylation at 'Thr-18' and 'Ser-20'. -
组织特异性
High expression is found in testis, spleen, colon and peripheral blood leukocytes. Low expression is found in other tissues. -
疾病相关
Defects in CHEK2 are associated with Li-Fraumeni syndrome 2 (LFS2) [MIM:609265]; a highly penetrant familial cancer phenotype usually associated with inherited mutations in p53/TP53.
Defects in CHEK2 may be a cause of susceptibility to prostate cancer (PC) [MIM:176807]. It is a malignancy originating in tissues of the prostate. Most prostate cancers are adenocarcinomas that develop in the acini of the prostatic ducts. Other rare histopathologic types of prostate cancer that occur in approximately 5% of patients include small cell carcinoma, mucinous carcinoma, prostatic ductal carcinoma, transitional cell carcinoma, squamous cell carcinoma, basal cell carcinoma, adenoid cystic carcinoma (basaloid), signet-ring cell carcinoma and neuroendocrine carcinoma.
Defects in CHEK2 are found in some patients with osteogenic sarcoma (OSRC) [MIM:259500]. -
序列相似性
Belongs to the protein kinase superfamily. CAMK Ser/Thr protein kinase family. CHK2 subfamily.
Contains 1 FHA domain.
Contains 1 protein kinase domain. -
翻译后修饰
Phosphorylated by PLK4. -
细胞定位
Nucleus; Nucleus. Isoform 10 is present throughout the cell and Nucleus > PML body. Nucleus > nucleoplasm. Recruited into PML bodies together with TP53. - Information by UniProt
-
数据库链接
- Entrez Gene: 11200 Human
- Omim: 604373 Human
- SwissProt: O96017 Human
- Unigene: 291363 Human
- Unigene: 505297 Human
-
别名
- CDS 1 antibody
- Cds1 antibody
- Cds1 homolog antibody
see all
图片
-
All lanes : Anti-Chk2 antibody [EPR4325] (ab109413) at 1/50000 dilution
Lane 1 : Wild-type A549 cell lysate
Lane 2 : CHEK2 knockout A549 cell lysate
Lane 3 : HEK-293 cell lysate
Lane 4 : MDA-MB-231 cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution
Performed under reducing conditions.
Predicted band size: 61 kDa
Observed band size: 67 kDa why is the actual band size different from the predicted?This data was developed using the same antibody clone in a different buffer formulation (ab109413)
False colour image of Western blot: Anti-Chk2 antibody [EPR4325] staining at 1/50000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab109413 was shown to bind specifically to Chk2. A band was observed at 67 kDa in wild-type A549 cell lysates with no signal observed at this size in CHEK2 knockout cell line ab276098 (knockout cell lysate ab276098).
To generate this image, wild-type and CHEK2 knockout A549 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.
-
All lanes : Anti-Chk2 antibody [EPR4325] (ab109413) at 1/1000 dilution
Lane 1 : Wild-type HeLa cell lysate
Lane 2 : CHEK2 knockout HeLa cell lysate
Lane 3 : HEK-293 cell lysate
Lane 4 : MDA-MB-231 cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 61 kDa
Observed band size: 68 kDa why is the actual band size different from the predicted?This data was developed using the same antibody clone in a different buffer formulation (ab109413).
Lanes 1-4: Merged signal (red and green). Green - ab109413 observed at 68 kDa. Red - loading control ab8245 observed at 37 kDa.
ab109413 Anti-Chk2 antibody [EPR4325] was shown to specifically react with Chk2 in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab264815 (knockout cell lysate ab257104) was used. Wild-type and Chk2 knockout samples were subjected to SDS-PAGE. ab109413 and Anti-alpha Tubulin antibody [EP1332Y] - Microtubule Marker (ab52866) were incubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
-
Intracellular Flow Cytometry analysis of HeLa (human cervix adenocarcinoma) cells labeling Chk2 with purified ab109413 at 1/230 dilution (10 µg/ml) (red). Cells were fixed with 80% methanol and permeabilised with 0.1% Tween-20. A Goat anti rabbit IgG (Alexa Fluor® 488) (ab150077) (1/2000 dilution) was used as the secondary antibody. Rabbit monoclonal IgG (Black) (ab172730) was used as the isotype control, cells without incubation with primary antibody and secondary antibody (Blue) were used as the unlabeled control.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab109413).
-
This data was developed using the same antibody clone in a different buffer formulation (ab109413).
Immunocytochemistry analysis of HT-29 (human colorectal adenocarcinoma epithelial cell) labeling Chk2 with purified ab109413 at 1/500 dilution. Cells were fixed with 4% Paraformaldehyde and permeabilised with 0.1% tritonX-100. Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) at 1/1000 (2 µg/ml) was used as the secondary antibody. was used as counterstain. Nuclei were stained blue with DAPI.
Negative control: PBS instead of the primary antibody. -
Immunohistochemical analysis of paraffin-embedded human spleen tissue using ab109413 at a 1/100 dilution.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab109413).
Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
-
This data was developed using ab109413, the same antibody clone in a different buffer formulation.
Purified ab109413 at 1/50 dilution (2µg) immunoprecipitating Chk2 in HeLa whole cell lysate.
Lane 1 (input): HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate 10µg
Lane 2 (+): ab109413 + HeLa whole cell lysate.
Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab109413 in HeLa whole cell lysate.
VeriBlot for IP Detection Reagent (HRP) (ab131366) (1/1000 dilution) was used for Western blotting.
Blocking Buffer and concentration: 5% NFDM/TBST.
Diluting buffer and concentration: 5% NFDM/TBST.
Observed band size: 62 kDa -
ab109413 staining Chk2 in wild-type HAP1 cells (top panel) and Chk2 knockout HAP1 cells (bottom panel). The cells were fixed with 100% methanol (5 minutes), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1 hour. The cells were then incubated with ab109413 at 1/250 dilution and ab195889 at 1/250 dilution (shown in pseudo colour red) overnight at +4°C, followed by a further incubation at room temperature for 1 hour with a goat secondary antibody to Rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green). Nuclear DNA was labelled in blue with DAPI.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab109413).
-
ab109413 (1/500) staining Chk2 in HeLa (human epithelial cell line from cervix adenocarcinoma) cells (green). Cells were fixed in paraformaldehyde, permeabilized with 0.5% Triton X-100/PBS and counterstained with DAPI in order to highlight the nucleus (red). For further experimental details please see Abreview.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab109413).
-
This ICC data was generated using the same anti-Chk2 antibody clone, EPR4325, in a different buffer formulation (cat# ab10413).
ab109413 staining Chk2 in wild-type HAP1 cells (top panel) and Chk2 knockout HAP1 cells (bottom panel). The cells were fixed with 100% methanol (5 minutes), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1 hour. The cells were then incubated with ab109413 at 1/250 dilution and ab195889 at 1/250 dilution (shown in pseudo colour red) overnight at +4°C, followed by a further incubation at room temperature for 1 hour with a goat secondary antibody to Rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green). Nuclear DNA was labelled in blue with DAPI.
-
This IHC data was generated using the same anti-Chk2 antibody clone, EPR4325, in a different buffer formulation (cat# ab109413).
Immunohistochemical analysis of paraffin-embedded human colon tissue using ab109413 at a 1/100 dilution.
Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
实验方案
To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.
数据表及文件
-
Datasheet download
Certificate of Compliance
文献 (4)
ab227998 被引用在 4 文献中.
- Tu Y et al. Ataxin-3 promotes genome integrity by stabilizing Chk1. Nucleic Acids Res 45:4532-4549 (2017). WB . PubMed: 28180282
- Li WF et al. WISP-1 contributes to fractionated irradiation-induced radioresistance in esophageal carcinoma cell lines and mice. PLoS One 9:e94751 (2014). Human . PubMed: 24728101
- Sowd GA et al. SV40 utilizes ATM kinase activity to prevent non-homologous end joining of broken viral DNA replication products. PLoS Pathog 10:e1004536 (2014). PubMed: 25474690
- Ogiwara H & Kohno T CBP and p300 histone acetyltransferases contribute to homologous recombination by transcriptionally activating the BRCA1 and RAD51 genes. PLoS One 7:e52810 (2012). PubMed: 23285190