Purpose: Our aim was to determine the repeatability of measurements of axial length (AL) and anterior chamber depth (ACD) made with two ultrasonic biometers and the IOLMaster in a group of non-orthokeratology (ortho-k) adult subjects and to investigate the agreement among instruments in children undergoing ortho-k therapy and in children wearing spectacles. Methods: To determine repeatability, AL and ACD were measured twice in 22 non-ortho-k young adults using two A-scan ultrasonic biometers (A-5500 and A-2500) and the IOLMaster. To determine agreement, AL and ACD were measured with the same instruments in 30 children undergoing ortho-k therapy and 30 spectacle-wearing children. Results: In the adult subjects, there were no significant differences in ACD and AL measurements obtained from the three instruments (repeated measures ANOVAs, p > 0.05). There was also no significant between-measurement difference for each instrument. The between-measurement agreement was better for the IOLMaster (95% limits of agreement (LA): -0.04 and +0.05 turn for both AL and ACD) than for the two A-scan ultrasonic biometers (95% LA: -0.12 and +0.11 turn for AL; -0.22 and +0.27 mm for ACD). Among the children, A-L measurements with all three instruments were not significantly different from each other for both the children undergoing ortho-k therapy and those wearing spectacles (repeated measures ANOVAs, p > 0.05). The 95% ILA of differences obtained from any two instruments were also comparable for both groups of subjects (within -0.20 mm and +0.20 mm). ACD measurements of the children were significantly different among the three instruments (repeated measures ANOVAs, p < 0.05). No significant differences in ACD measurements were found between A-5500 and A-2500 for both groups of children (paired t tests, p > 0.017). Conclusions: The repeatability of AL and ACD measurements with the IOLMaster was very good, and was better than with the A-scan ultrasonic biometers. The agreements in AL measurements between A-scan ultrasonic biometers; and IOLMaster were comparable in both the ortho-k and the spectacle-wearing subjects, and were comparable to the repeatability of the A-scan ultrasonic biometers. ACD measurements between A-scan ultrasonic biometry and the IOLMaster were not comparable. AL measurements with the IOLMaster can replace the measurements from the two A-scan ultrasonic biometers used, however, the reverse is not true. AL and ACD measurements with all three instruments were unaffected by the flattened cornea following ortho-k lens wear.