Fig. 5. Localisation of active LFA-1 and cathepsin X in the migrating T lymphocytes. Jurkat T lymphocytes were seeded on ICAM1 pre-coated slides and allowed to migrate for 15 minutes before labelling. Active LFA-1 was detected with Alexa_fluor-488-conjugated specific antibody mAb 24. (A) Wild-type Jurkat T lymphocytes. Active LFA-1 is localised manly in the perimembranous region of the spherical non-migratory state cells. Cathepsin-X-overexpressing Jurkat T lymphocytes showing polarised phenotype. Active LFA-1 is localised predominantly at the uropod region (white arrows). (B) Colocalisation of active LFA-1 and cathepsin X in upregulated migratory cells along the z axis. Active LFA-1 is localised at the mid-cell region, which is in contact with ICAM1-coated surface (0.0 µm) and at the uropod projecting above the surface (2.7 µm). (C) Colocalisation of active cathepsin X (green; Alexa-Fluor-488) and the lysosomal marker protein LAMP2 (red; Alexa-Fluor-546) in cathepsin-X-overexpressing T lymphocytes. Cathepsin X is colocalised in the lysosomes (yellow), it is also present in perimembrane region and uropod (green). Scale bars, 50 µm (A), 5 µm (B), 10 µm (C).