The HDAC Activity Assay Kit is designed to measure HDAC activity in cell or nuclear extracts, immunoprecipitates or purified enzymes.
Catalogue Number
566328
Brand Family
Calbiochem®
Application Data
1Refers to dilution of Trichostatin A in HDAC Assay Buffer, which will be 5X the final concentration. Examples: 1) As a measure of non-HDAC background, 5 µM would produce final 1 µM concentration and essentially complete HDAC inhibition; 2) As a model inhibitor "hit", 25 nM would produce final 5 nM and ~50% inhibition. 2 Refers to dilution of potential inhibitor in Assay Buffer, which will be 5x its final concentration. 1The appropriate dilution of the Deacetylated Standard may be determined from the standard curve and should be the concentration producing a fluorescent signal equal to that produced by control (no inhibitor) samples in the HDAC assay. The dilution in HDAC Assay Buffer is prepared at 1.25X this concentration to compensate for the 4/5 dilution due to addition of 10 µl of Assay Buffer or inhibitor. 2 Refers to dilution of Trichostatin A in HDAC Assay Buffer, which will be 5X its final concentration in the 50 µl volume, prior to addition of HDAC Developer WS. Example: As a model inhibitor that does not interfere with the HDAC Developer, 25 nM Trichostatin A would produce a final 5 nM concentration. 3Refers to dilution of potential inhibitor in HDAC Assay Buffer, which will be 5X its final concentration in the 50 µl volume, prior to addition of HDAC Developer WS. 50 µl of each diluted Deacetylated Standard was mixed with 50 µl HDAC Developer and incubated for 10 min at 25°C. Fluorescence was then measured using the clear 1/2 Volume Plate with a fluorimeter (PerSeptive Biosystems, Ex. 360 nm, Em. 460 nm, gain=85)
Materials Required but Not Delivered
• Fluorimeter for measuring fluorescence in a 96-well plate at an excitation wavelength of 350-380 nm and an emission wavelength of 440-460 nm. • Pipetman or multi-channel pipetman capable of pipetting 2-100 µl accurately • Ice bucket to keep reagents cold until use. • Plate warmer or other temperature control device (optional)
References
References
Gurvich, N., et al. 2004. Cancer Res.64, 1079. Kapustin, G.V., et al. 2003. Org. Lett.5, 3053. Bitterman, K.J., et al. 2002. J. Biol. Chem.277, 45099. Grozinger, C.M., et al. 2001. J. Biol. Chem.276, 38837.
Product Information
Form
96 Tests
Format
96-well plate
Applications
Biological Information
Sample Type
cell extracts, immunoprecipitates, or purified enzymes
Physicochemical Information
Dimensions
Materials Information
Toxicological Information
Safety Information according to GHS
Safety Information
R Phrase
R: 36/38
Irritating to eyes and skin.
S Phrase
S: 26-36-45
In case of contact with eyes, rinse immediately with plenty of water and seek medical advice. Wear suitable protective clothing. In case of accident or if you feel unwell, seek medical advice immediately (show the label where possible).
Product Usage Statements
Intended use
The Calbiochem® HDAC Activity assay is a complete assay system designed to measure histone deacetylase (HDAC) activity in cell or nuclear extracts, immunoprecipitates, or purified enzymes. It has been used successfully with preparations of all class I HDACs-HDAC1, HDAC2, HDAC3 and HDAC8 - with class II HDACs 4-7, 9 and 10 and with yeast Sir2 and its human homolog, SIRT1.
Storage and Shipping Information
Ship Code
Dry Ice Only
Toxicity
Multiple Toxicity Values, refer to MSDS
Storage
≤ -70°C
Storage Conditions
Upon arrival store the HeLa Cell Nuclear Extract, HDAC Substrate, HDAC Developer Concentrate, Trichostatin A (HDAC Inhibitor), Deacetylated Standard, and HDAC Assay Buffer at -70°C. Store both of the 1/2 Volume Microplates at room temperature.
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