Depth of interaction (DOI) information is indispensable to improving the sensitivity and spatial resolution of positron emission tomography (PET) systems, especially for small field-of-view PET such as small animal PET and human brain PET. We have already developed a series of X’tal cube detectors for isotropic spatial resolution and we obtained the best isotropic resolution of 0.77 mm for detectors with six-sided readout. However, it is still challenging to apply the detector for PET systems due to the high cost of six-sided readout electronics and carrying out segmentation of a monolithic cubic scintillator in three dimensions using the subsurface laser engraving (SSLE) technique. In this work, we propose a more practical X’tal cube with a two-sided readout detector, which is made of crystal bars segmented in the height direction only by using the SSLE technique. We developed two types of prototype detectors with a 3 mm cubic segment and a 1.5 mm cubic segment by using 3 × 3 × 20 mm3 and 1.5 × 1.5 × 20 mm3 crystal bars segmented into 7 and 13 DOI segments, respectively, using the SSLE technique. First, the performance of the detector, composed of one crystal bar with different DOI segments and two thorough silicon via (TSV) multi-pixel photon counters (MPPCs) as readout at both ends of the crystal bar, were evaluated in order to demonstrate the capability of the segmented crystal bars as a DOI detector. Then, performance evaluation was carried out for a 4 × 4 crystal array of 3 × 3 × 20 mm3 with 7 DOI segments and an 8 × 8 crystal array of 1.5 × 1.5 × 20 mm3 with 13 DOI segments. Each readout included a 4 × 4 channel of the 3 × 3 mm2 active area of the TSV MPPCs. The three-dimensional position maps of the detectors were obtained by the Anger-type calculation. All the segments in the 4 × 4 array were identified very clearly when there was air between the crystal bars, as each crystal bar was coupled to one channel of the MPPCs; however, it was necessary to optimize optical conditions between crystal bars for the 8 × 8 array because of light sharing between crystal bars coupled to one channel of the MPPCs. The optimization was performed for the 8 × 8 array by inserting reflectors fully or partially between the crystal bars and the best crystal identification performance was obtained with the partial reflectors between the crystal bars. The mean energy resolutions at the 511 keV photo peak for the 4 × 4 array with air between the crystal bars and for the 8 × 8 array with partial reflectors between the crystal bars were 10.1% ± 0.3% and 10.8% ± 0.8%, respectively. Timing resolutions of 783 ± 36 ps and 1.14 ± 0.22 ns were obtained for the detectors composed of the 4 × 4 array and the 8 × 8 array with partial reflectors, respectively. These values correspond to single photon timing resolutions. Practical X’tal cubes with 3 mm and 1.5 mm DOI resolutions and two-sided readout were developed.