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Caspase-3 DEVD-R110 Fluorometric HTS Assay Kit

A fluorometric endpoint assay kit for detection of caspase-3 activity in cells by high throughput screening (HTS).

Product Attributes

Apoptosis/viability marker

Caspase

For live or fixed cells

Cell lysis required

Detection method/readout

Microplate reader (fluorescence)

Assay type/options

Endpoint assay, Homogeneous assay, High-throughput assay

Substrate specificity

Caspases

Colors

Green

Excitation/Emission

496/520 nm (end product)

Storage Conditions

Store at -10 to -35 °C

Size
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10 assays
100 assays
1,000 assays
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Protocols
SDS

Product Description

Caspase-3 DEVD-R110 Fluorometric HTS Assay Kit  is a homogenous assay system for fast and highly sensitive detection of caspase-3 activity in mammalian cells. The kit is specifically designed for high throughput screening (HTS)-based assays. For real-time detection of caspase-3 activity in intact cells, learn about our novel NucView® Caspase-3 Substrates.

Features

  • Caspase-3 activity measured with substrate (Ac-DEVD)2-R110
  • Designed for high throughout assays
  • Fast enzyme kinetics
  • Enzymatic reaction forms green fluorescent R110 product
  • Endpoint assay requires cell lysis

Kit Components

  • Cell lysis/assay buffer
  • Enzyme substrate (Ac-DEVD)2-R110
  • Enzyme inhibitor Ac-DEVD-CHO
  • R110

About this kit

Caspase-3 is an active cell-death protease involved in the execution phase of apoptosis. The fluorogenic and chromogenic substrate (Ac-DEVD)2-R110 contains two DEVD tetrapeptides and is completely hydrolyzed by the caspase-3 in two successive steps. Cleavage of the first DEVD peptide results in the monopeptide Ac-DEVD-R110 intermediate, which has absorption and emission wavelengths similar to those of R110 (rhodamine 110) (Ex/Em= 496/520 nm) but has only about 10% of the fluorescence of the latter. Hydrolysis of the second DEVD peptide releases the dye R110, leading to a substantial fluorescence increase.

The assay kit includes Ac-DEVD-CHO, which is a caspase-3 inhibitor and can be used as a negative control. Also, R110 is provided in the kit for generating a standard curve, which can be used for quantifying caspase-3 activity.

Note: While caspase-3 preferentially cleaves the consensus sequence DEVD compared to other substrate sequences, other caspases can also cleave DEVD efficiently. Overlapping caspase substrate recognition limits the usefulness of caspase substrate peptides for distinguishing between different caspase activities in cell lysates.

NucView® enzyme substrate technology is covered by U.S. patents.

References

1. World J Gastroenterol (2008) Jan 7; 14(1): 76–80 doi: 10.3748/wjg.14.76
2. Biochim Biophys Acta (2015) Jul;1851(7):919-28 doi: 10.1016/j.bbalip.2015.03.001
3. Indian J Med Res (2015) 141, pp807-815 doi:10.4103/0971-5916.160713
4. J Autoimmun (2015) Jun;60:40-50 doi: 10.1016/j.jaut.2015.03.005.
5. J Clin Invest (2016) Sep 1;126(9):3336-50 doi: 10.1172/JCI85939.
6. J Lipid Res (2016) Jul;57(7):1231-42 doi: 10.1194/jlr.M067389.
7. Oncotarget (2016) Nov 22; 7(47): 77696–77706 doi:10.18632/oncotarget.12773

 

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