In this study, a new radio-frequency (RF)-based, three-dimensional (3-D) strain imaging technique is introduced and applied to 3-D full volume ultrasound data of the heart of healthy children. Continuing advances in performance of transducers for 3-D ultrasound imaging have boosted research on 3-D strain imaging. In general, speckle tracking techniques are used for strain imaging. RF-based strain imaging has the potential to yield better performance than speckle- based methods because of the availability of phase information but such a system output is commercially not available. Furthermore, the relatively low frame rate of 3-D ultrasound data has limited broad application of RF-based cardiac strain imaging. In this study, the previously reported two-dimensional (2-D) strain methodology was extended to the third dimension. Three-dimensional RF-data were acquired in 13 healthy children, in the age range of 6-15 years, at a relatively low frame rate of 38-51 Hz. A 3-D, free-shape, coarse-to-fine displacement and strain estimation algorithm was applied to the RF-data. The heart was segmented using 3-D ellipsoid fitting. Strain was estimated in the radial (R), circumferential (C) and longitudinal directions (L). Our preliminary results reveal the applicability of the 3-D strain estimation technique on full volume 3-D RF-data. The technique enabled 3-D strain imaging of all three strain components. The average strains for all children were in the lateral wall R = 37 � 10\% (infero-lateral) and R = 32\% � 10\% (antero-lateral), C = -9\% � 4\% (antero-lateral) and C = -9\% � 4\% (infero-lateral), L = -18\% � 6 \% (antero-lateral) and L = -15\% � 4\% (infero-lateral). In the septum, strains were found to be R = 24\% � 10\% (antero-septal) and R = 13\% � 5\% (infero-septal), C = -13\% � 5\% (antero-septal) and -13\% � 5\% (infero-septal) and L = -13\% � 3\% (antero-septal) and L = -16\% � 5\% (infero-septal). Strain in the anterior and inferior walls seemed underestimated, probably caused by the low (in-plane) resolution and poor image quality. The field-of-view as well as image quality were not always sufficient to image the entire left ventricle. It is concluded that 3-D strain imaging using RF-data is feasible, but validation with other modalities and with conventional 3-D speckle tracking techniques will be necessary.