Anti-phospho-Chk1(Ser345) Antibody, clone 3D14 ZooMAb® Rabbit Monoclonal
SIGMA/ZRB1424 - recombinant, expressed in HEK 293 cells
Product Type: Chemical
Immunocytochemistry Enhanced Validation - Recombinant Antibody Technology Immunofluorescent analysis of untreated HeLa cells (A and B) and HeLa cells treated with UV light (50 milliJoules with 1 hr recovery) (C and D) were performed using a 1:100 dilution of Cat. No. ZRB1424, Anti-phospho-Chk1(Ser345), clone 3D14 ZooMAb® Rabbit Monoclonal and visualized with a Goat Anti-Rabbit secondary antibody conjugated to Alexa Fluor® 488 (Green). Actin filaments have been labeled with phalloidin (Red). Nucleus is stained with DAPI (Blue). This antibody positively stains the nucleus.
Immunocytochemistry Enhanced Validation - Recombinant Antibody Technology Immunofluorescent analysis of untreated HeLa cells (A and B) and HeLa cells treated with UV light (50 milliJoules with 1 hr recovery) (C and D) were performed using a 1:100 dilution of Cat. No. ZRB1424, Anti-phospho-Chk1(Ser345), clone 3D14 ZooMAb® Rabbit Monoclonal and visualized with a Goat Anti-Rabbit secondary antibody conjugated to Alexa Fluor® 488 (Green). Actin filaments have been labeled with phalloidin (Red). Nucleus is stained with DAPI (Blue). This antibody positively stains the nucleus.
Immunocytochemistry Enhanced Validation - Recombinant Antibody Technology Immunofluorescent analysis of untreated HeLa cells (A and B) and HeLa cells treated with UV light (50 milliJoules with 1 hr recovery) (C and D) were performed using a 1:100 dilution of Cat. No. ZRB1424, Anti-phospho-Chk1(Ser345), clone 3D14 ZooMAb® Rabbit Monoclonal and visualized with a Goat Anti-Rabbit secondary antibody conjugated to Alexa Fluor® 488 (Green). Actin filaments have been labeled with phalloidin (Red). Nucleus is stained with DAPI (Blue). This antibody positively stains the nucleus.
Immunocytochemistry Enhanced Validation - Recombinant Antibody Technology Immunofluorescent analysis of untreated HeLa cells (A and B) and HeLa cells treated with UV light (50 milliJoules with 1 hr recovery) (C and D) were performed using a 1:100 dilution of Cat. No. ZRB1424, Anti-phospho-Chk1(Ser345), clone 3D14 ZooMAb® Rabbit Monoclonal and visualized with a Goat Anti-Rabbit secondary antibody conjugated to Alexa Fluor® 488 (Green). Actin filaments have been labeled with phalloidin (Red). Nucleus is stained with DAPI (Blue). This antibody positively stains the nucleus.
Immunocytochemistry Enhanced Validation - Recombinant Antibody Technology Immunofluorescent analysis of untreated HeLa cells (A and B) and HeLa cells treated with UV light (50 milliJoules with 1 hr recovery) (C and D) were performed using a 1:100 dilution of Cat. No. ZRB1424, Anti-phospho-Chk1(Ser345), clone 3D14 ZooMAb® Rabbit Monoclonal and visualized with a Goat Anti-Rabbit secondary antibody conjugated to Alexa Fluor® 488 (Green). Actin filaments have been labeled with phalloidin (Red). Nucleus is stained with DAPI (Blue). This antibody positively stains the nucleus.
Western Blotting Enhanced Validation - Recombinant Antibody Technology Lysates from untreated HeLa cells (Lane 1) and HeLa treated with Etoposide (20 μg/mL) (Lane 2) were probed with Cat. No. ZRB1424, Anti-phospho-Chk1(Ser345), clone 3D14 ZooMAb® Rabbit Monoclonal (1:1,000 dilution). Proteins were visualized using a Donkey Anti-Rabbit IgG secondary antibody conjugated to HRP and a chemiluminescence detection system. Arrow indicates phospho-Chk1(Ser345) (~55 kDa).
Affinity Binding Assay Enhanced Validation - Recombinant Antibody Technology 100 mg/mL of non-specific control peptide (Run 1) or Phospho-Chk1(Ser435) peptide (Run 2) were analyzed by affinity binding assay using Cat. No. ZRB1424, Anti-phospho-Chk1 (Ser345), clone 3D14 ZooMAb® Rabbit Monoclonal. The binding of this antibody to Phospho-Chk1-Ser435 peptide displayed a KD of 3.0 x 10-6, whereas binding to non-specific control peptide displayed a KD of 2.3 x 10-3.
12 Principles of Green Chemistry: Principle 1—Waste Prevention. This product has a smaller environmental footprint through reduction of waste in the manufacturing process.
12 Principles of Green Chemistry: Principle 4—Designing Safer Chemicals
Chemical products should be designed to affect their desired function while minimizing their toxicity.
Chemical products should be designed to affect their desired function while minimizing their toxicity.
12 Principles of Green Chemistry: Principle 6—Design for Energy Efficiency. This product was re-engineered to utilize less energy in the manufacturing process.
| antibody form | purified antibody |
| antibody product type | primary antibodies |
| biological source | rabbit |
| clone | 3D14, recombinant monoclonal |
| conjugate | unconjugated |
| description | recombinant, expressed in HEK 293 cells |
| enhanced validation | recombinant expression Learn more about Antibody Enhanced Validation  |
| epitope sequence | C-terminal half |
| form | lyophilized |
| greener alternative category | Aligned |
| greener alternative product characteristics | Waste Prevention Designing Safer Chemicals Design for Energy Efficiency Learn more about the Principles of Green Chemistry . |
| isotype | IgG |
| mol wt | calculated mol wt 54.43 kDa |
| observed mol wt ~55 kDa | |
| packaging | antibody small pack of 25 μL |
| product line | ZooMAb® learn more |
| Protein ID accession no. | NP_001107593  |
| purified by | using Protein A |
| Quality Level | 200 |
| recombinant | expressed in HEK 293 cells |
| shipped in | ambient |
| species reactivity | monkey, mouse, human |
| species reactivity (predicted by homology) | canine, porcine, feline |
| storage temp. | 2-8°C |
| technique(s) | affinity binding assay: suitable |
| immunocytochemistry: suitable | |
| western blot: suitable | |
| UniProt accession no. | O14757 |
| Application: | Quality Control Testing Evaluated by Western Blotting in lysate from HeLa cells treated with Etoposide. Western Blotting Analysis: A 1:1,000 dilution of this antibody detected Serine/Threonine-protein kinase Chk1 phosphorylated on Ser345 in lysate from HeLa cells overnight serum starved and treated with Etoposide (20 μg/ml), but not in lysate from untreated cells. Tested applications Western Blotting Analysis: A 1:1,000 dilution from a representative lot detected phospho-Chk1 (Ser345) in COS-1 and NIH3T3 cells treated with UV lighrt. Immunocytochemistry Analysis: A 1:100 dilution from a representative lot detected phospho-Chk1 (Ser345) in HeLa cells treated with UV light (50 mJ UV w/1hr recovery). Affinity Binding Assay: A representative lot of this antibody bound phosphorylated Chk1 with a KD of 3.0 x 10-6 in an affinity binding assay. Note: Actual optimal working dilutions must be determined by end user as specimens, and experimental conditions may vary with the end user |
| Application: | Anti-phospho-Chk1 (Ser345), clone 3D14 ZooMAb®, Cat. No. ZRB1424, is a recombinant Rabbit Monoclonal antibody that detects Chk1 phosphorylated on Ser345 and is tested in Affinity Binding Assay, Immunocytochemistry, and Western Blotting. |
| Disclaimer: | Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals. |
| General description: | We are committed to bringing you greener alternative products, which adhere to one or more of The 12 Principles of Green Chemistry.This antibody is Preservative-free, produced without the harm or sacrifice of animals and exceptionally stable to allow for ambient shipping and storage if needed and thus aligns with "Waste Prevention", "Designing Safer Chemicals" and "Design for Energy Efficiency". Click here for more information. |
| General description: | ZooMAb® antibodies represent an entirely new generation of recombinant monoclonal antibodies.Each ZooMAb® antibody is manufactured using our proprietary recombinant expression system, purified to homogeneity, and precisely dispensed to produce robust and highly reproducible lot-to-lot consistency. Only top-performing clones are released for use by researchers. Each antibody is validated for high specificity and affinity across multiple applications, including its most commonly used application. ZooMAb® antibodies are reliably available and ready to ship when you need them. |
| Immunogen: | KLH-conjugated linear peptide corresponding to 11 amino acids surrounding phosphoserine 345 from the C-terminal half of human Serine/threonine-protein kinase Chk1 |
| Legal Information: | ZooMAb is a registered trademark of Merck KGaA, Darmstadt, Germany |
| Physical form: | Purified recombinant rabbit monoclonal antibody IgG, lyophilized in PBS with 5% Trehalose, normal appearance a coarse or translucent resin. The PBS/trehalose components in the ZooMAb® formulation can have the appearance of a semi-solid (bead like gel) after lyophilization. This is a normal phenomenon. Please follow the recommended reconstitution procedure in the data sheet to dissolve the semi-solid, bead-like, gel-appearing material. The resulting antibody solution is completely stable and functional as proven by full functional testing. Contains no biocide or preservatives, such as azide, or any animal by-products. Larger pack sizes provided as multiples of 25 μL. |
| Reconstitution: | 300 μg/mL after reconstitution at 25 μL per vial. Please refer to guidance on suggested starting dilutions and/or titers per application and sample type. |
| Specificity: | Clone 3D14 is a ZooMAb® Rabbit recombinant monoclonal antibody that specifically detects Serine/threonine-protein kinase Chk1 phosphorylated on serine 345. |
| Storage and Stability: | Recommend storage of lyophilized product at 2-8°C; Before reconstitution, micro-centrifuge vials briefly to spin down material to bottom of the vial; Reconstitute each vial by adding 25 μL of filtered lab grade water or PBS; Reconstituted antibodies can be stored at 2-8°C, or -20°C for long term storage. Avoid repeated freeze-thaws. |
| Target description: | Serine/threonine-protein kinase Chk1 (UniProt: O14757; also known as EC:2.7.11.1, CHK1 checkpoint homolog, Cell cycle checkpoint kinase, Checkpoint kinase-1) is encoded by the CHEK1 (also known as CHK1) gene (Gene ID: 1111) in human. Chk1 is a serine/threonine-protein kinase that is required for checkpoint-mediated cell cycle arrest. It is expressed ubiquitously, and its most abundant expression is observed in thymus, testis, small intestine, and colon. It is activated through phosphorylation predominantly by ATR but also by ATM in response to DNA damage or inhibition of DNA replication. It may also negatively regulate cell cycle progression during unperturbed cell cycles that is achieved by a number of mechanisms that together help to preserve the integrity of the genome. Chk1 recognizes the substrate consensus sequence [R-X-X-S/T]. It binds to and phosphorylates CDC25A (at Ser 178, Thr 507) and CDC25C (at Ser 216). These phosphorylations create binding sies for 14-3-3- proteins, which inhibit CDC2A and CDC2C. Phosphorylation of Chk1 on serine 345 induces a change in the conformation of the protein that activates the kinase activity and is shown to be a prerequisite for interaction with FBXO6 and subsequent ubiquitination at lysine 436. Phosphorylation at serine 345 increases its binding to 14-3-3 proteins and promotes its nuclear retention. Conversely, dephosphorylation at serine 345 by protein phosphatase, Mg2+/Mn2+ dependent 1D (PPM1D) contributes to exit from checkpoint mediated cell cycle arrest. The activated, serine 345 phosphorylated form can undergo polyubiquitination at lysine 436 and undergo degradation ensuring that activated Chk1 does not accumulate as cells progress through S phase. This ZooMAb® recombinant monoclonal antibody, generated by our propriety technology, offers significantly enhanced specificity, affinity, reproducibility, and stability over conventional monoclonals. Ref.: Moiseeva, TN., et al. (2019). Proc. Natl. Acad. Sci. USA. 116(27); 13374-13383; Chen, MS., et al. (2003). Mol. Cell Biol. 23(21); 7488-7497). |
| Storage Temp. | 2-8°C |