A protocol for visualizing active cathepsin K in osteoclasts with a quenched-fluorescence-activity-based probe.
STAR Protoc
; 4(3): 102465, 2023 Sep 15.
Article
em En
| MEDLINE
| ID: mdl-37481728
ABSTRACT
Herein, we provide a protocol for visualizing active osteoclast cathepsin K (CatK) with the quenched-fluorescent-activity-based probe qTJK17. We describe steps for isolating peripheral blood mononuclear cells, their differentiation into osteoclasts, and TRAP staining using an acid phosphatase leukocyte kit. We then detail visualization of active CatK. The probe qTJK17 includes a reactive group, acyloxymethylketone, that binds to the CatK active site, recognition sequence, and fluorescence donor-acceptor pair. This protocol can determine the exact localization of active CatK in osteoclasts. For complete details on the use and execution of this protocol, please refer to Janiszewski et al. (2023).1.
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Base de dados:
MEDLINE
Assunto principal:
Osteoclastos
/
Corantes Fluorescentes
Idioma:
En
Ano de publicação:
2023
Tipo de documento:
Article