Fig. 1. Fluorokine E enzyme activity assay schematic.
Fluorokine E enzyme activity assays utilize a fluorogenic substrate linked to a quencher molecule. This enzyme detection system facilitates the measurement of active MMPs without the use of zymography (see Figure 1). MMP present within the sample is captured by a specific antibody pre-coated onto a black microplate. The active MMP cleaves the peptide between the fluorophore and the quencher molecule eliminating the distance dependent resonance energy transfer. This produces a fluorescent signal proportional to the amount of active enzyme present in the sample.
An activation reagent, APMA, is added to all standards and selected samples. The kit is designed to measure endogenous, naturally occurring active MMP present in samples and any forms that can be activated by the addition of APMA.
Levels of total active MMP (endogenous and forms that can be activated) can be compared to naturally occurring MMP by assaying the same sample with and without APMA.