SensoLyte® Homogeneous Rh110 Caspase-3/7 Assay Kit *Fluorimetric*

500 Assays (96-well plate)

Central to the execution phase of apoptosis are the two closely related caspase-3 and caspase-7. They share common substrate specificity and structure, but differ completely in the sequence of their respective N-terminal regions. Both caspase-3 and caspase-7 have substrate selectivity for the amino acid sequence Asp-Glu-Val-Asp (DEVD).
The SensoLyte® Homogeneous Rh110 Capase-3/7 Assay Kit uses (Ac-DEVD)2-Rh110 as the fluorogenic indicator for assaying caspase-3/7 activities. Upon caspase-3/7 cleavage, (Ac-DEVD)2-Rh110 generates the Rh110 fluorophore which has bright green fluorescence and can be detected at Ex/Em=496 nm/520 nm. The longer-wavelength spectra and higher extinction coefficient of the Rh110 provide greater sensitivity and less interference from cell components.

A bi-function assay buffer in this kit is designed to lyze the cells and measure the enzyme activity at the same time. Thus, this kit can measure caspase-3/7 activity in cell culture directly in a 96-well or 384-well plate without a time-consuming cell extraction step. In case the cells are cultured in larger plates or flasks, a lysis buffer and protocol for cell lysate preparation are also conveniently included in the kit. The kit is suitable for high throughput screening of apoptosis inducers and inhibitors.

Kits components    
Component A: Rh110 Caspase-3/7 substrate Peptide sequence=(Z-Asp-Glu-Val-Asp)2-Rh110, Ex/Em=496 nm/520 nm upon caspase-3/7 cleavage: 250 µL Component B: Rh110, fluorescence reference standard, Ex/Em=496 nm/520 nm: 1 mM DMSO solution, 40 µL Component C: Ac-DEVD-CHO, a known caspase-3/7 inhibitor Peptide sequence=Ac-Asp-Glu-Val-Asp-CHO: 5 mM DMSO solution, 10 µL Component D: Assay buffe:r 30 mL Component E: DTT: 1 M, 1.1 mL Component F: 10X Lysis Buffer: 20 mL


Absorbance (nm)- 496
Emission (nm)- 520
    
Store components A, B, C, and E at -20°C. Components D and F can be stored at 4°C for convenience. Component A and B protect from light and moisture.

Application    
- FRET
- Protease activity

Biomarker Target    
- Caspase
- Caspase 3
- Caspase 7

Detection Method    
-Fluorescent

Research Area    
- Cancer & Apoptosis