In order to assess the accuracy and reliability of age estimates from calcified structures in the three-spined stickleback Gasterosteus aculeatus, we evaluated intra and inter-reader repeatability from three structures: otoliths, gill covers and pelvic spines). Average age estimates were also compared between the structures. The overall intra-reader repeatability of age estimates were highest for otoliths (69%), lowest for gill covers (53%) and intermediate for spine cross-sections (63%). Although four of the seven readers had the highest intra-reader repeatability score for spine cross-sections, the inter-reader variance in this structure was much higher than in others. Otoliths were the easiest in terms of their pre-analysis treatment and exchange of materials (as digital images) between readers. In addition, otoliths are more well-studied compared with the other structures with respect to their development through ontogenesis; hence, age estimates based on otoliths should be the most reliable. Therefore, our recommendation is that whenever possible, analysis of otoliths should be the preferred approach for aging G. aculeatus.