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HDAC Activity Colorimetric Assay Kit

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Antigen
  • ARABIDOPSIS HISTONE DEACETYLASE 1
  • ARABIDOPSIS HISTONE DEACETYLASE 19
  • ATHD1
  • ATHDA19
  • ATRPD3A
  • F20D10.250
  • F20D10_250
  • HDA1
  • HDA19
  • HISTONE DEACETYLASE
  • HISTONE DEACETYLASE 19
  • HISTONE DEACETYLASE19
  • RPD3A
  • histone deacetylase 1
  • ARABIDOPSIS HISTONE DEACETYLASE 2
  • ATHD2
  • ATHD2B
  • HD2
  • HDA4
  • HDT02
  • HDT2
  • HISTONE DEACETYLASE 2
  • MDJ22.7
  • MDJ22_7
  • histone deacetylase 2B
  • Tb05.26K5.290
  • Histone DeAcetylase
  • Protein HDA-4
  • histone deacetylase 19
  • histone deacetylase HDT2
  • histone deacetylase
  • Histone deacetylase
  • hda-3
  • hda-6
  • hda-1
  • hda-4
  • HD1
  • HD2B
  • Tb927.2.2190
  • Tb927.5.2900
  • PAS_chr1-4_0125
  • PAS_chr2-2_0016
Reactivity
Mammalian
4
2
2
1
1
1
1
1
1
1
1
1
1
1
Application
Activity Assay (AcA), High-throughput Screening (HTS)
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Sample Type Nuclear Extract
Detection Method Colorimetric
Specificity The new Colorimetric HDAC Activity Assay Kit provides a fast and convenient colorimetric method that eliminates radioactivity, extractions, or chromatography, as used in the traditional assays. The new method requires only two easy steps, both performed on the same microtiter plate. first, the HDAC colorimetric substrate, which comprises an acetylated lysine side chain, is incubated with a sample containing HDAC activity (e.g., HeLa nuclear extract or your own samples). Deacetylation of the substrate sensitizes the substrate, so that, in the second step, treatment with the Lysine Developer produces a chromophore. The chromophore can be easily analyzed using an ELISA plate reader or spectrophotometer. The assay is well suited for high throughput screening applications. HDAC inhibitors and antibodies are also available separately
Characteristics Colorimetric HDAC Activity Assay Kit: Measure Histone Deacetylase (HDAC) Activity in Just Two Easy Steps. Convenient, High-Throughput Adaptable Colorimetric Assay.
Components HDAC Substrate [Ac-Lys(Ac)-pNA, 10 mM]
10X HDAC Assay Buffer
Lysine Developer
HDAC Inhibitor (Trichostatin A, 1 mM)
HeLa Nuclear Extract (5 mg/ml)
Deacetylated Standard (Ac-Lys-pNA, 10 mM)
Alternative Name HDAC (HDAC ELISA Kit Abstract)
Background Inhibition of histone deacetylases (HDACs) has been implicated to modulate transcription and to induce apoptosis or differentiation in cancer cells. However, screening HDAC inhibitory compounds has proven to be difficult over the past due to the lack of convenient tools for analyzing HDAC activity. The new Colorimetric HDAC Activity Assay Kit provides a fast and convenient colorimetric method that eliminates radioactivity, extractions, or chromatography, as used in the traditional assays. The new method requires only two easy steps, both performed on the same microtiter plate. First, the HDAC colorimetric substrate, which comprises an acetylated lysine side chain, is incubated with a sample containing HDAC activity (e.g., HeLa nuclear extract or your own samples). Deacetylation of the substrate sensitizes the substrate, so that, in the second step, treatment with the Lysine Developer produces a chromophore. The chromophore can be easily analyzed using an ELISA plate reader or spectrophotometer. The assay is well suited for high throughput screening applications. HDAC inhibitors and antibodies are also available separately.
Application Notes Well suited for high throughput applications. HDAC inhibitors and substrates and related antibodies are also available separately.
Comment

Further details regarding sample type: Cell lysate, nuclear extract

Protocol A. General Consideration: Read the entire protocol before beginning the procedure. The HeLa nuclear extract and Lysine Developer should be refreeze immediately at -20 or -70 °C after each use to avoid loss of activity. If positive and negative controls are designed, the kit provides sufficient reagents for 5 positive control assays with the HeLa Nuclear Extract and 5 Negative Control assays with the HDAC Inhibitor, Trichostatin A. Using 96-well plates with U-shape bottom. Flat bottom may give a little low value. B.
1. Dilute test samples (50-200 µg of nuclear extract or cell lysate) to 85 µL (final volume) of ddH 2 0 in each well (For background reading, add 85 µL ddH 2 0 only). For positive control, dilute 10 µL of HeLa nuclear extract with 75 µL ddH 2 0. For negative control, dilute yoursample into 83 µL of ddH 2 0 and then add 2 µL of Trichostatin, or use a known sample containing no HDAC activity.
2. Add 10 µL of the 10X HDAC Assay Buffer to each well.
3. Add 5 µL of the HDAC colorimetric substrate to each well. Mix thoroughly.
4. Incubate plates at 37 °C for 1 hour (or longer if desired).
5. Stop the reaction by adding 10 µL of Lysine Developer and mix well. Incubate the plate at 37 °C for 30 min.
6. Read sample in an ELISA plate reader at 400 or 405 nm. Signal is stable for several hours at room temperature. HDAC activity can be expressed as the relative O.D. value per µg protein sample. C. Standard Curve (optional):
1. If desired, a standard curve can be prepared using the known amount of the Deacetylated Standard included in the kit. The exact concentration range of the Deacetylase Standard will vary depending on the each individual plate reader and the exact wavelength used. We recommend starting with a dilution range of 10-100 µM in Assay Buffer.
2. Add 90 µL each of the dilutions and also 10 µL of the 10X Assay Buffer into a set of wells on the microtiter plate. Use 90 µL of H2O and 10 µL of 10X Assay Buffer as zero
3. Add 10 µL of Lysine Developer to each well and incubate at 37 °C for 30 min (Note: Incubation time should be kept the same for both standard and test samples.)
4. Read samples in an ELISA plate reader at 400 or 405 nm.
5. Plot O.D. value (y-axis) versus concentration of the Deacetylated Standard (x-axis). Determine the slope as delta O.D./ µM.
6. Based on the slope, you can determine the absolute amount of deacetylated lysine generated in your sample.
Restrictions For Research Use only
Storage -20 °C
Expiry Date 12 months
Product cited in: Yang, Li, Chen, Yuan, Dong, Zhang, Wu, Wang: "PARP-1 mediates LPS-induced HMGB1 release by macrophages through regulation of HMGB1 acetylation." in: Journal of immunology (Baltimore, Md. : 1950), Vol. 193, Issue 12, pp. 6114-23, 2014 (PubMed).

Zhang, DeNicola, Qin, Du, Ma, Tina Zhao, Zhuang, Liu, Wei, Qin, Tang, Zhao: "Specific inhibition of HDAC4 in cardiac progenitor cells enhances myocardial repairs." in: American journal of physiology. Cell physiology, Vol. 307, Issue 4, pp. C358-72, 2014 (PubMed).

Aslan, Phillips, Healy, Itakura, Pang, McCarty: "Histone deacetylase 6-mediated deacetylation of α-tubulin coordinates cytoskeletal and signaling events during platelet activation." in: American journal of physiology. Cell physiology, Vol. 305, Issue 12, pp. C1230-9, 2013 (PubMed).

Spallotta, Cencioni, Straino, Nanni, Rosati, Artuso, Manni, Colussi, Piaggio, Martelli, Valente, Mai, Capogrossi, Farsetti, Gaetano: "A nitric oxide-dependent cross-talk between class I and III histone deacetylases accelerates skin repair." in: The Journal of biological chemistry, Vol. 288, Issue 16, pp. 11004-12, 2013 (PubMed).

Alsadany, Shehata, Mohamad, Mahfouz: "Histone deacetylases enzyme, copper, and IL-8 levels in patients with Alzheimer's disease." in: American journal of Alzheimer's disease and other dementias, Vol. 28, Issue 1, pp. 54-61, 2013 (PubMed).

Gnana-Prakasam, Veeranan-Karmegam, Coothankandaswamy, Reddy, Martin, Thangaraju, Smith, Ganapathy: "Loss of Hfe leads to progression of tumor phenotype in primary retinal pigment epithelial cells." in: Investigative ophthalmology & visual science, Vol. 54, Issue 1, pp. 63-71, 2013 (PubMed).

Wu, Zheng, Huang, Yan, Yin, Xu, Zhang, Gui, Chu, Liu: "Lithium down-regulates histone deacetylase 1 (HDAC1) and induces degradation of mutant huntingtin." in: The Journal of biological chemistry, Vol. 288, Issue 49, pp. 35500-10, 2013 (PubMed).

Candelaria, de la Cruz-Hernandez, Taja-Chayeb, Perez-Cardenas, Trejo-Becerril, Gonzalez-Fierro, Chavez-Blanco, Soto-Reyes, Dominguez, Trujillo, Diaz-Chavez, Duenas-Gonzalez: "DNA methylation-independent reversion of gemcitabine resistance by hydralazine in cervical cancer cells." in: PLoS ONE, Vol. 7, Issue 3, pp. e29181, 2012 (PubMed).

Zhang, Qin, Zhao, Fast, Zhuang, Liu, Cheng, Zhao: "Inhibition of histone deacetylases preserves myocardial performance and prevents cardiac remodeling through stimulation of endogenous angiomyogenesis." in: The Journal of pharmacology and experimental therapeutics, Vol. 341, Issue 1, pp. 285-93, 2012 (PubMed).

Dioum, Osborne, Goetsch, Russell, Schneider, Cobb: "A small molecule differentiation inducer increases insulin production by pancreatic ? cells." in: Proceedings of the National Academy of Sciences of the United States of America, Vol. 108, Issue 51, pp. 20713-8, 2011 (PubMed).

Minetti, Feige, Rosenstiel, Bombard, Meier, Werner, Bassilana, Sailer, Kahle, Lambert, Glass, Fornaro: "Gαi2 signaling promotes skeletal muscle hypertrophy, myoblast differentiation, and muscle regeneration." in: Science signaling, Vol. 4, Issue 201, pp. ra80, 2011 (PubMed).

Mizuno, Yasuo, Bogaard, Kraskauskas, Natarajan, Voelkel: "Inhibition of histone deacetylase causes emphysema." in: American journal of physiology. Lung cellular and molecular physiology, Vol. 300, Issue 3, pp. L402-13, 2011 (PubMed).

Carter, Lin, Liu, Yang, Liu: "Phosphorylated p68 RNA helicase activates Snail1 transcription by promoting HDAC1 dissociation from the Snail1 promoter." in: Oncogene, Vol. 29, Issue 39, pp. 5427-36, 2010 (PubMed).

Roccaro, Sacco, Jia, Azab, Maiso, Ngo, Azab, Runnels, Quang, Ghobrial: "microRNA-dependent modulation of histone acetylation in Waldenstrom macroglobulinemia." in: Blood, Vol. 116, Issue 9, pp. 1506-14, 2010 (PubMed).

Singh, Thangaraju, Prasad, Martin, Lambert, Boettger, Offermanns, Ganapathy: "Blockade of dendritic cell development by bacterial fermentation products butyrate and propionate through a transporter (Slc5a8)-dependent inhibition of histone deacetylases." in: The Journal of biological chemistry, Vol. 285, Issue 36, pp. 27601-8, 2010 (PubMed).

Zhou, Deng, Norseen, Lieberman: "Regulation of Epstein-Barr virus origin of plasmid replication (OriP) by the S-phase checkpoint kinase Chk2." in: Journal of virology, Vol. 84, Issue 10, pp. 4979-87, 2010 (PubMed).

Choi, Jung, Lee, Yoon, Kwon, Kang, Kim, Cha, Kim, Jun, Lee, Yoon: "Epigallocatechin-3-gallate, a histone acetyltransferase inhibitor, inhibits EBV-induced B lymphocyte transformation via suppression of RelA acetylation." in: Cancer research, Vol. 69, Issue 2, pp. 583-92, 2009 (PubMed).

Farhana, Dawson, Dannenberg, Xu, Fontana: "SHP and Sin3A expression are essential for adamantyl-substituted retinoid-related molecule-mediated nuclear factor-kappaB activation, c-Fos/c-Jun expression, and cellular apoptosis." in: Molecular cancer therapeutics, Vol. 8, Issue 6, pp. 1625-35, 2009 (PubMed).

Bourguignon, Xia, Wong et al.: "Hyaluronan-mediated CD44 interaction with p300 and SIRT1 regulates beta-catenin signaling and NFkappaB-specific transcription activity leading to MDR1 and Bcl-xL gene expression and chemoresistance ..." in: The Journal of biological chemistry, Vol. 284, Issue 5, pp. 2657-71, 2009 (PubMed).

Liu, Mai, Sun: "Lysine acetylation regulates Bruton's tyrosine kinase in B cell activation." in: Journal of immunology (Baltimore, Md. : 1950), Vol. 184, Issue 1, pp. 244-54, 2009 (PubMed).