Mk podosomes are longer-lived on collagen. (A) Lifeact-GFP cells were spread on 100 μg/mL fibrinogen (top images) or 100 μg/mL Horm collagen-coated surfaces (bottom images) and were imaged in real time with a TIRF microscope for 20 minutes (fibrinogen) and 2 hours (Horm collagen) using a 100× objective. Pictures were taken every 10 seconds (fibrinogen) or 20 seconds (Horm collagen). Red squares show enlarged single podosomes. (B) The average lifetime ±SEM was determined by measuring 100 podosomes in total of 10 different cells. Data were collected of 4 independent experiments. (C) Cumulative frequency analysis of podosome lifetime was completed in 2-minute intervals. A Student t test was used for the statistical analysis. Asterisk indicates significant difference with a P value < .05. Error bars indicate ±SEM. See also supplemental Movie 1 (podosomes on fibrinogen), supplemental Movie 2 (podosomes on collagen), and supplemental Movie 3 (WASp in podosomes on collagen).