Purpose: A clinically applicable and accurate method for measuring Achilles tendon length is needed to investigate the influence of elongation of the Achilles tendon after acute rupture. The purpose of this study was to develop and validate an ultrasonographic (US) length measurement of the Achilles tendon-aponeurosis complex. Methods: Both legs of 19 non-injured subjects were examined by magnetic resonance imaging (MRI) and US. The length from calcaneus to the medial head of m. Gastrocnemius was measured by three independent US examiners. Repeated US measurements were performed and compared to MRI measurements. Intra-rater and inter-rater reliability and the agreement between MRI and US were determined. Data were evaluated using the intraclass correlation coefficient (ICC), the standard error of the measurement (SEM) and the minimal detectable change (MDC). Results: Intra-rater reliability of US assessment showed no significant differences between test days: ICC 0.96, SEM 4 mm and MDC 10 mm. Inter-rater reliability showed a systematic difference between US observers of 2–5 mm (p = 0.001–0.036); ICC 0.97, SEM 3 mm and MDC 9 mm. MRI measurements were on average 4 mm longer than US (p = 0.001). Conclusion: The novel ultrasound measurement showed good reliability and accuracy. For comparison between groups of non-injured subjects differences of more than 4 mm can be detected. For repeated assessment of individual subjects differences of more than 10 mm can be detected. The measurement needs to be further assessed in the setting of acute Achilles tendon rupture. Clinical relevance: This new ultrasound measurement might allow for length measurement of ruptured Achilles tendons in the acute and chronic state after rupture. Level of evidence: II.