Anti-phospho-HDAC4 (S246) Antibody, clone 1I14 ZooMAb® Rabbit Monoclonal
SIGMA/ZRB1547 - recombinant, expressed in HEK 293 cells
Synonym: Histone deacetylase 4;EC:3.5.1.98;HD4
Product Type: Chemical
Immunohistochemistry (Paraffin) Enhanced Validation - Recombinant Antibody Technology Formalin Fixed Paraffin Embedded (FFPE) human breast cancer (A) and Negative control (no primary) (B) tissue sections were prepared using heat-induced epitope retrieval (HIER). Immunostaining was performed using a 1:100 dilution of Cat. No. ZRB1547, Anti-phospho-HDAC4 (S246), clone 1I14 ZooMAb® Rabbit Monoclonal. Reactivity was detected using a Goat Anti-Rabbit IgG and HRP-DAB. Cytoplasmic/membranous/nuclear staining was observed in human breast cancer tissue sections.
Immunohistochemistry (Paraffin) Enhanced Validation - Recombinant Antibody Technology Formalin Fixed Paraffin Embedded (FFPE) human breast cancer (A) and Negative control (no primary) (B) tissue sections were prepared using heat-induced epitope retrieval (HIER). Immunostaining was performed using a 1:100 dilution of Cat. No. ZRB1547, Anti-phospho-HDAC4 (S246), clone 1I14 ZooMAb® Rabbit Monoclonal. Reactivity was detected using a Goat Anti-Rabbit IgG and HRP-DAB. Cytoplasmic/membranous/nuclear staining was observed in human breast cancer tissue sections.
Immunohistochemistry (Paraffin) Enhanced Validation - Recombinant Antibody Technology Formalin Fixed Paraffin Embedded (FFPE) human breast cancer (A) and Negative control (no primary) (B) tissue sections were prepared using heat-induced epitope retrieval (HIER). Immunostaining was performed using a 1:100 dilution of Cat. No. ZRB1547, Anti-phospho-HDAC4 (S246), clone 1I14 ZooMAb® Rabbit Monoclonal. Reactivity was detected using a Goat Anti-Rabbit IgG and HRP-DAB. Cytoplasmic/membranous/nuclear staining was observed in human breast cancer tissue sections.
Immunocytochemistry Enhanced Validation - Recombinant Antibody Technology Immunofluorescent analysis of Untreated HEK293 cells (A and B) and HEK293 treated with human recombinant EGF (C and D) was performed using a 1:100 dilution of Cat. No. ZRB1547, Anti-phospho-HDAC4 (S246), clone 1I14 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 HEK293 cells (A and B) and HEK293 treated with human recombinant EGF (C and D) was performed using a 1:100 dilution of Cat. No. ZRB1547, Anti-phospho-HDAC4 (S246), clone 1I14 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 HEK293 cells (A and B) and HEK293 treated with human recombinant EGF (C and D) was performed using a 1:100 dilution of Cat. No. ZRB1547, Anti-phospho-HDAC4 (S246), clone 1I14 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 HEK293 cells (A and B) and HEK293 treated with human recombinant EGF (C and D) was performed using a 1:100 dilution of Cat. No. ZRB1547, Anti-phospho-HDAC4 (S246), clone 1I14 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 HEK293 cells (A and B) and HEK293 treated with human recombinant EGF (C and D) was performed using a 1:100 dilution of Cat. No. ZRB1547, Anti-phospho-HDAC4 (S246), clone 1I14 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 Lysate from PC12 cells treated with human recombinant EGF (25 ng/mL for 15 min) was probed with Cat. No. ZRB1547, Anti-phospho-HDAC4 (S246), clone 1I14 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-HDAC4 (S246) (~110 kDa).
Western Blotting Enhanced Validation - Recombinant Antibody Technology Lysates from overnight serum starved, untreated HEK293 cells (Lane 1) and HEK293 cells treated with human recombinant EGF (50 ng/mL for 10 min) (Lane 2) were probed with 1:1,000 dilution of Cat. No. ZRB1547, Anti-phospho-HDAC4 (S246), clone 1I14 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-HDAC4 (S246) (~110 and 120 kDa).
Affinity Binding Assay Enhanced Validation - Recombinant Antibody Technology 100 mg/mL of non-phosphorylated HDAC4 peptide (Run 1) or phosphorylated HDAC4 (Ser246) peptide (Run 2) were analyzed by affinity binding assay using Cat. No. ZRB1547, Anti-phospho-HDAC4 (S246), clone 1I14 ZooMAb® Rabbit Monoclonal. The binding of this antibody to phosphorylated HDAC4 peptide displayed a KD of 7.8 x 10-8, whereas binding to non-phosphorylated HDAC4 control peptide displayed a KD of 1.7 x 10-5.
Peptide Inhibition Assay Enhanced Validation - Recombinant Antibody Technology Lysates from HEK293 cells treated with human recombinant EGF was probed with Cat. No. ZRB1547, Anti-phospho-HDAC4 (S246), clone 1I14 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-HDAC4 (S246) (~110 and 120 kDa). Lanes 1. Lysate from HEK293 cells without any peptide treatment. 2. Lysate from HEK293 cells treated with human recombinant EGF and incubated with phosphorylated HDAC4 (Ser246) peptide. 3. Lysate from HEK293 cells treated with human recombinant EGF and incubated with a non-phosphorylated HDAC4 peptide.
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 | 1I14, recombinant monoclonal |
| conjugate | unconjugated |
| description | recombinant, expressed in HEK 293 cells |
| enhanced validation | recombinant expression Learn more about Antibody Enhanced Validation  |
| epitope sequence | N-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 119.04 and 106.37 kDa |
| observed mol wt ~120 and 110 kDa | |
| packaging | antibody small pack of 25 μL |
| product line | ZooMAb® learn more |
| Protein ID accession no. | NP_006028  |
| purified by | using Protein A |
| Quality Level | 200 |
| recombinant | expressed in HEK 293 cells |
| shipped in | ambient |
| species reactivity | human, rat |
| species reactivity (predicted by homology) | bovine, monkey, canine, feline, mouse |
| storage temp. | 2-8°C |
| technique(s) | affinity binding assay: suitable |
| immunocytochemistry: suitable | |
| immunohistochemistry (formalin-fixed, paraffin-embedded sections): suitable | |
| western blot: suitable | |
| UniProt accession no. | P56524 |
| Application: | Quality Control Testing Evaluated by Western Blotting in lysate from HEK293 cells treated with EGF. Western Blotting Analysis (WB): A 1:1,000 dilution of this antibody detected HDAC4 phosphorylated on serine 246 in lysate from overnight serum starved HEK293 cells treated with recombinant human EGF (50 ng/mL for 10 min). Tested applications Western Blotting Analysis: A 1:1,000 dilution from a representative lot detected phospho-HDAC4 (S246) in PC12 treated with EGF. Affinity Binding Assay: A representative lot of this antibody bound phospho-HDAC4 (Ser246) with a KD of 7.8 x 10-8 in an affinity binding assay. Immunohistochemistry (Paraffin) Analysis: A 1:100 dilution from a representative lot detected phospho-HDAC4 (S246) in human breast cancer tissue sections. Immunocytochemistry Analysis: A 1:100 dilution from a representative lot detected phospho-HDAC4 (S246) in HEK293 cells treated with EGF. Peptide Inhibition Assay: Target band detection in lysate from HEK293 cells treated with EGF was prevented by preblocking of a representative lot with phospho-HDAC4 peptide. Note: Actual optimal working dilutions must be determined by end user as specimens, and experimental conditions may vary with the end user |
| 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 10 amino acids surrounding phosphoserine 246 from the N-terminal half of human HDAC4. |
| Legal Information: | ZooMAb is a registered trademark of Merck KGaA, Darmstadt, Germany |
| Physical form: | Purified recombinant rabbit monoclonal antibody IgG, lyophilized in PBS, 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 1I14 is a ZooMAb® Rabbit recombinant monoclonal antibody that specifically detects Histone deacetylase 4 (HDAC4) phosphorylated on serine 246. It detects both isoforms of HDAC4. |
| 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: | Histone deacetylase 4 (UniProt: P56524; also known as EC:3.5.1.98, HD4) is encoded by the HDAC4 (also known as KIAA0288) gene (Gene ID: 9759) in human. HDAC4 is a ubiquitous enzyme that is responsible for the deacetylation of lysine residues on the N-terminal part of the core histones (H2A, H2B, H3 and H4). Histone deacetylation gives a tag for epigenetic repression and plays an important role in transcriptional regulation, cell cycle progression, and developmental events. It is present as a homodimeric protein that shuttles between the nucleus and the cytoplasm. Its nuclear export sequence is localized in amino acids 1051-1084. Its export to cytoplasm depends on the interaction with a 14-3-3 chaperone protein. 14-3-3 family of proteins are known to bind specifically to conserved phosphoserine-containing motifs. Phosphorylation of specific amino acids creates binding sites for the 14-3-3, which escorts phospho- HDAC4 from the nucleus to the cytoplasm. Phosphorylation of HDAC4 at serine 246, 467, and 632 is achieved by CaMK4. Its nuclear localization probably depends on sumoylation on lysine 559, which is promoted by E3 SUMO-protein ligase RanBP2. Two isoforms of HDAC4 have been described that are produced by alternative splicing. Isoform 2 lacks the first 117 amino acids. This ZooMAb® recombinant monoclonal antibody, generated by our propriety technology, offers significantly enhanced specificity, affinity, reproducibility, and stability over conventional monoclonals. (Ref.: Wang, Z., et al. (2014). Epigenomics 6(1); 139-150; Wang, AH., et al. (2000). Mol. Cell Biol. 20(18); 6904-6912). |
| WGK Germany | WGK 1 |
| Flash Point(F) | Not applicable |
| Flash Point(C) | Not applicable |
| Storage Temp. | 2-8°C |