Studies have indicated that adding 2-D quasi-static elastography to B-mode ultrasound imaging improved the specificity for malignant lesion detection, as malignant lesions are often stiffer (increased strain ratio) compared with benign lesions. This method is limited by its user dependency and so unsuitable for breast screening. To overcome this limitation, we implemented quasi-static elastography in an automated breast volume scanner (ABVS), which is an operator-independent 3-D ultrasound system and is especially useful for screening women with dense breasts. The study aim was to investigate if 3-D quasi-static elastography implemented in a clinically used ABVS can discriminate between benign and malignant breast lesions. Volumetric breast ultrasound radiofrequency data sets of 82 patients were acquired before and after automated transducer lifting. Lesions were annotated and strain was calculated using an in-house-developed strain algorithm. Two strain ratio types were calculated per lesion: using axial and maximal principal strain (i.e., strain in dominant direction). Forty-four lesions were detected: 9 carcinomas, 23 cysts and 12 other benign lesions. A significant difference was found between malignant (median: 1.7, range: [1.0-3.2]) and benign (1.0, [0.6-1.9]) using maximal principal strain ratios. Axial strain ratio did not reveal a significant difference between benign (0.6, [-12.7 to 4.9]) and malignant lesions (0.8, [-3.5 to 5.1]). Three-dimensional strain imaging was successfully implemented on a clinically used ABVS to obtain, visualize and analyze in vivo strain images in three dimensions. Results revealed that maximal principal strain ratios are significantly increased in malignant compared with benign lesions.