Fig. 5. Analysis of the role of gelsolin in actin depolymerisation and insulin secretion by exogenous expression of an HA-tagged gelsolin cDNA construct in B1 and C3 cells. (A) Western blot showing the effect of exogenous expression of HA-gelsolin in B1 and C3 cells on the total level of gelsolin. Exogenous HA-gelsolin was specifically detected with an anti-HA-tag antibody (10 µg total protein/lane). (B) Immunofluorescence analysis of the subcellular localisation of HA-gelsolin (green) and the actin cytoskeleton (red) on B1 (top panels) and C3 cells (bottom panels) with or without 20 mM glucose stimulation. Nuclei (blue) were stained with DRAQ5^TM. For glucose-stimulated B1 cells, two examples of HA-gelsolin-expressing cells are given and the actin (red) signal is boosted in the higher magnification image (zoom) to facilitate visualisation of actin/gelsolin co-localisation. Bars, 10 µm (unless stated otherwise). (C) Growth hormone secretion by B1 cells co-transfected with an HA-gelsolin expression plasmid or with a control empty plasmid. Data are mean ± s.e.m. of three replicates of a single experiment.