Lysosomes are a cell organelle type processing antimicrobial activity. Here, we investigate the lysosomal activity in a primary cell line, bovine aortic endothelial cells (BAECs), during cellular aging, based on the antimicrobial activity of lysosomes isolated from BAECs at cell passages 4, 6, 8, and 10. Cytochemical analysis of lysosomes with LysoTracker reagent revealed the number of lysosome-like organelles surrounding the nucleus initially increased drastically in the BAECs and continued increasing gradually until passage 10. The lysosomes isolated from each successive passage of BAECs exhibited increased antimicrobial activity against Escherichia coli, and, in addition, an age-dependent increase in lysosome intensity coincided with increased lysosomal antimicrobial activity.