Figure 1
From: Cystatin E/M suppresses legumain activity and invasion of human melanoma
Cystatin E/M and C secretions from various cell lines: Common laboratory non-melanoma and established laboratory melanoma cell lines (top), and primary and metastatic melanoma cell lines established from patients, as well as skin control (bottom). (A) Equal amounts of serum free media were collected from 5 × 105 cells 48 h after changing from ordinary growth media and secreted proteins were concentrated by TCA-precipitation and subjected to SDS-PAGE and immunoblotting. The filters were stained with a cystatin E/M-specific (upper panels) or a cystatin C-specific (lower panel) antibody, respectively. (B) Inhibitory activity against legumain was measured in the conditioned media as residual legumain activity. A partially purified legumain fraction from rat kidney was mixed with conditioned media and the ability to cleave the substrate Z-Ala-Ala-Asn-NHMec was measured by fluorometry. Control bar (100%) represents activity in the rat legumain fraction without addition of conditioned media.