Cathepsins are apoptosis markers associated with Alzheimer′s disease, numerous types of cancer, autoimmune arthritis, and the breakdown of bone structure seen with osteoporosis.
Cathepsin K detection assay uses (z-Leucine-Arginine)2(z-LR)2 derivative of the cresyl violet fluorophore, which easily penetrates the cell membrane and the membranes of the internal cellular organelles enabling detection of cathepsin K activity within whole living cells. Hoechst stain is used to label the cell nuclei after labeling with the MR- Cathepsin K substrate. It is visualized under a microscope using a UV-filter with excitation at 365 nm and emission at 480 nm. Acridine orange (AO) stains aggregates in acidic pH and fluoreses orange rather than green, thus clearly differentiating the lysosomes from the other organelles.
Cathepsin B, K, and L protease activities can be detected within whole living cells using substrate-based fluorescent assays, which utilize cathepsin target sequence peptides, derivatives of the cresyl violet fluorophore, conjugated to Magic Red™ (MR) fluorogenic substrate (cresyl violet). Following enzymatic cleavage inside the living cells at one or both arginine (R) amide linkage sites, the mono and non-substituted cresyl violet fluorophores generate red fluorescence when excited at 550-590 nm.