Western blot analysis of prostasin expression in Calu-3 sublines
Top panel: total cell lysate, 40 µg/lane; middle panel: Triton™ X-114-extracted membrane fractions from 40 µg of total cell lysate; bottom panel: isolated exosomes from 0.5 ml conditioned medium per sample (2.5 × 105 cells). (A) Samples were immunoblotted with an antibody against human prostasin. (B) Samples were immunoblotted with antibodies against β-tubulin (total lysate) or CD63 (membrane fractions and exosomes) as loading controls. Calu: Calu-3 parent cells, CC: Calu-3 control subline with scrambled gRNA, KO: Calu-3 knockout subline with prostasin-specific gRNA, V: Calu-3 subline with vector alone, P: Calu-3 subline overexpressing the wildtype prostasin, M: Calu-3 subline overexpressing an active-site mutant prostasin, G: Calu-3 overexpressing an active prostasin without the GPI-anchor.