Our website does not fully support your browser.

We've detected that you are using an older version of Internet Explorer. Your commerce experience may be limited. Please update your browser to Internet Explorer 11 or above.

We believe this site might serve you best:

United States

United States

Language: English

Promega's Cookie Policy

Our website uses functional cookies that do not collect any personal information or track your browsing activity. When you select your country, you agree that we can place these functional cookies on your device.

ApoTox-Glo™ Triplex Assay Technical Manual

Instructions for Use of Product(s)
G6320, G6321

Literature # TM322

The ApoTox-Glo™ Triplex Assay combines three Promega assay chemistries to assess viability, cytotoxicity and caspase activation events within a single assay well. The first part of the assay simultaneously measures two protease activities; one is a marker of cell viability, and the other is a marker of cytotoxicity. The live-cell protease activity is restricted to intact viable cells and is measured using a fluorogenic, cell-permeant, peptide substrate (GF‑AFC). The substrate enters intact cells where it is cleaved by the live-cell protease activity to generate a fluorescent signal proportional to the number of living cells. This live-cell protease becomes inactive upon loss of cell membrane integrity and leakage into the surrounding culture medium. A second, fluorogenic cell-impermeant peptide substrate (bis-AAF-R110) is used to measure dead-cell protease activity, which is released from cells that have lost membrane integrity. Because bis-AAF-R110 is not cell-permeant, essentially no signal from this substrate is generated by intact, viable cells. The live- and dead-cell proteases produce different products, AFC and R110, which have different excitation and emission spectra, allowing them to be detected simultaneously.

The second part of the assay uses a luminogenic caspase-3/7 substrate, which contains the tetrapeptide sequence DEVD, in a reagent optimized for caspase activity, luciferase activity and cell lysis. Adding the Caspase-Glo® 3/7 Reagent in an “add-mix-measure” format results in cell lysis, followed by caspase cleavage of the substrate and generation of a “glow-type” luminescent signal produced by luciferase. Luminescence is proportional to the amount of caspase activity present.

Summary of Changes
The following changes were made to the 4/15 revision of this document:
1. The patent/license statements were updated.
2. The document design was updated.

Printed in USA. Revised 4/15.

Experienced User Protocols