Related Experiment Video
Updated: Jun 7, 2025
Non-invasive In Vivo Fluorescence Optical Imaging of Inflammatory MMP Activity Using an Activatable Fluorescent Imaging Agent
Published on: May 8, 2017
Environment-sensitive turn-on fluorescent probe enables live cell imaging of myeloperoxidase activity during NETosis
Enebie Ramos Cáceres1, Lotte Kemperman1, Kimberly M Bonger2
1Department of Synthetic Organic Chemistry, Institute for Molecules and Materials, Radboud University, Heyendaalseweg 135, 6525AJ, Nijmegen, The Netherlands.
Abstract:
Myeloperoxidase (MPO) plays an important role in the immune response of human neutrophils and has been implicated in autoimmune conditions, cardiovascular disorders, and neurodegeneration. Current methods to detect MPO activity rely on the detection of HOCl using activatable probes or require challenging experimental procedures. Therefore, these tools provide limited information about the dynamics and localization of MPO in complex molecular processes such as NETosis in real time. In this study, we report a ''turn-on" activity-based probe that fluoresces exclusively upon binding to MPO, exhibits minimal background fluorescence in buffered aqueous media, and is blocked by MPO inhibitors. Our probe facilitates real-time imaging of direct MPO activity in human neutrophils and HL-60-derived granulocytes during NETosis under wash-free conditions. Furthermore, it allows for the discrimination between different triggers of NETosis in human neutrophils. These findings hold promise for advancing our understanding of the role of MPO in immune responses and inflammatory conditions.
Insights
Researchers developed a new fluorescent probe to track myeloperoxidase (MPO) activity in real-time during neutrophil immune responses like NETosis. This tool offers better insights into MPO
Area of Science:
- Biochemistry
- Immunology
- Cell Biology
Background:
- Myeloperoxidase (MPO) is crucial in neutrophil immune responses and linked to various diseases.
- Existing MPO detection methods are limited for real-time dynamics and localization studies.
- Assessing MPO activity during neutrophil extracellular trap formation (NETosis) is challenging.
Purpose of the Study:
- To develop and validate a novel "turn-on" activity-based probe for MPO.
- To enable real-time imaging of MPO activity in neutrophils during NETosis.
- To differentiate NETosis triggers based on MPO activity.
Main Methods:
- Design and synthesis of a "turn-on" fluorescent probe specific for MPO.
- Testing probe specificity, background fluorescence, and inhibition by MPO inhibitors.
- Real-time imaging of MPO activity in human neutrophils and HL-60 cells during NETosis.
- Utilizing the probe under wash-free conditions.
Main Results:
- The developed probe exhibits high specificity for MPO with minimal background fluorescence.
- The probe successfully visualizes direct MPO activity in real-time during NETosis.
- Wash-free imaging conditions were achieved.
- The probe differentiated between various NETosis triggers in human neutrophils.
Conclusions:
- The novel activity-based probe allows for unprecedented real-time monitoring of MPO activity.
- This tool enhances understanding of MPO's role in neutrophil functions and inflammatory diseases.
- The probe facilitates discrimination of NETosis pathways, aiding future research.
