User Manual

Predefined methods
Eppendorf
®
PlateReader AF2200
English (EN)
67
6.2.8 Cell viability (Apo-ONE Caspase-3/7 Assay)
Introduction
The Apo-ONE Homogeneous Caspase-3/7 Assay provides a homogenous fluorometric method for
monitoring apoptosis in mammalian cells. The assay is based on the measurement of the activities of the
proteases caspase-3 and -7 in the process of apoptosis. A pro-fluorescent substrate becomes fluorescent
after cleavage by the activated caspases.
Default measurement parameters
• Excitation wavelength: 485 (25) nm
• Emission wavelength: 535 (35) nm
•Number of flashes: 25
• Settle time: 0 ms
•Mode: Top
• Lag time: 0 μs
• Integration: 20 μs
• Gain: "optimal"
• Plate definition: Eppendorf Cell Culture Plate 96/F, black/clear" or comparable plates
User defined method parameters
• The measurement is performed at 485 / 535 nm with default measurement parameters.
• Define the number of blank replicates (possible number: 1 – 6).
• Define the number of control replicates (possible number: 1 – 6).
• Define number of samples and sample replicates (max. 58, depends on number of blanks and controls).
• Define appropriate sample identifier (default settings are ID_1, ID_2, ..).
• Check if entered parameters are correct.
• Required plate layout: Outer wells are left empty by default. Pipette in a vertical order, starting with
blanks at position B2, controls are pipetted in subsequent wells, samples must be placed in the
following wells.
• Press Start.
• For method results and data evaluation principle see chapter method results and evaluation procedure.
The method is restricted to 96-well microplates.
Because of faster evaporation the wells lying on the outer sides of the plate are not included in
the evaluation.
 Please fill them with buffer or distilled water.