Sample Files

Caspase-3 AMC Substrate

November 21, 2012

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Related Products: Gen5 for Detection


Gen5 version required: 1.0 or higher


Basis for the Assay:

Caspase proteins are proteolytic enzymes involved in programmed cell death or Apoptosis. This proteolytic processing occurs at critical aspartic acid residues that conform to the caspase recognition sequence. Different caspase enzymes recognize related but different protein sequences. Caspase-3 can be monitored using a synthetic peptide, which releases a fluorometric moiety when cleaved by the enzyme. Prior to cleavage the substrate is nonfluorescent. The increase in fluorescence is proportional to the amount of caspase-3 activity present in the sample.

The protocol calls for a fluorescence measurement using a 360/40 excitation filter and a 460/40 emission filter.

The experiment data file contains all of the fluorescence determinations for a Caspase-3 enzyme activity determination using a peptide substrate with an AMC fluorescent moiety.

Plate Configuration:

Plate configuration assumes that samples will be run in duplicate. Included is a standard curve of Caspase-3 enzyme activities (0 to 1.6 ng/well). The plate includes 8 standard concentrations and two blank wells, but no controls. Thus, 40 samples can be run on a microplate in duplicate, in addition to the standard curve.

Report:

The report is configured to provide the (1) raw fluorescence, (2) blanked fluorescence, (3) standard curve, and (4) calculated caspase-3 concentrations of unknown samples.

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