One step detection of matrix metalloproteinase activity using a fluorogenic peptide probe-immobilized diagnostic kit
- One step detection of matrix metalloproteinase activity using a fluorogenic peptide probe-immobilized diagnostic kit
- 류주희; 이애주; 이슬기; 안철희; 박종웅; James F. Leary; 박상진; 김광명; 권익찬; 윤인찬; 최귀원
- Issue Date
- Bioconjugate chemistry
- VOL 21, 1378-1384
- Matrix metalloproteinases (MMPs) have been shown to be abundant in pathological conditions such as cancer,
osteoarthritis (OA), and rheumatoid arthritis (RA). The extent of MMPs detected in biological samples provides
important clinical information for diagnosis, prognosis, and therapeutic monitoring of various diseases relating
with MMPs. Herein, we developed a new high-throughput MMP diagnostic kit (MMP-D-KIT) based on a 96-
well plate by immobilizing MMP-13 specific fluorogenic peptide probes (MMP peptide probe), which is a pair
consisting of a near-infrared (NIR) fluorophore (Cy5.5) and a quencher (BHQ-3), onto the biocompatible glycol
chitosan (GC) polymer anchored 96-well plate. When MMP enzymes were simply added and incubated in a
MMP-D-KIT, the fluorescence of each well was recovered and the fluorescence intensity showed distinct difference
within minutes through NIR fluorescence imaging system. The fluorescence was recovered not only by MMP-13
activity, but also by other MMPs activity. Furthermore, recovery of NIR fluorescent signals in MMP-D-KIT was
proportional to concentrations of immobilized MMP peptide probe-GC conjugates and, importantly, MMP
concentration. The MMP-D-KIT is most specific for target MMP, compared with other enzymes including caspase-3
and 20s proteasome. Additionally, the MMP-D-KIT was used to detect MMP activity in biological samples such
as synovial fluid from 12 OA patients (grades 1-4 based on the Kellgren-Lawrence grading scale). It was found
that the fluorescence intensity measured using MMP-D-KIT decidedly correlates with the progression of OA.
The MMP-D-KIT could be applicable in detecting MMP activities in various biological samples and evaluating
the effects of MMP inhibitors in a rapid and easy fashion.
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