Hoechst stain can be used to label the cell nuclei after labeling with the MR-Cathepsin B substrate. It is visualized under a microscope using a UV-filter with excitation at 365 nm and emission at 480 nm. Acridine orange (AO) helps identify lysosomes and other intracellular organelles. In the acidic pH of the lysosome AO, molecules aggregate. Aggregated AO fluoreses orange rather than green thus clearly differentiating the lysosomes from the other organelles.
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 B kit detects protease activity within whole living cells as a marker of intracellular cathepsin B activity. In the assay (z-Arginine-Arginine)2 (z-RR)2 derivative of the cresyl violet fluorophore easily penetrates the cell membrane and the membranes of the internal cellular organelles enabling to detect cathepsin B activity within whole living cells.
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.