The purpose of this study was to assess the effects of fluence, pulse stacking, and multiple passes on the depth of injury caused by a fractionated Er:YAG laser in an in vivo farm pig model. DESIGN/MATERIAL/METHODS: A fractionated 2,940 nm Er:YAG laser (Pixel, Alma Lasers, Caesarea, Israel) was applied to the flank skin of a Yorkshire cross pig. The 11 mmx11 mm handpiece was comprised of either 49 or 81 microbeams (200 microm diameter), depending on the tip configuration. There were six different parameter sets divided according to total energy per pulse (150, 285, and 500 mJ) and tip type (81 or 49 microbeams per 11 mmx11 mm macrospot). Each of these six groups was subdivided according to number of stacked pulses (1, 3, and 6) and number of passes (1, 3, and 6). This resulted in a total of 36 treatment parameters. With the 49 microbeam configuration, a single pulse resulted in partial epidermal ablation at 150 mJ, complete epidermal ablation at 285 mJ and partial dermal ablation at 500 mJ to a depth of 90 microm. Stacking the pulses resulted in a significant increase in ablation with each fluence with the maximal depth of ablation measured at 140 microm after six stacked pulses at 500 mJ. Increasing the number of passes did not result in a significant increase in ablative depth, but did create a larger surface area of ablation. Residual thermal damage (RTD) was minimal and remained between 10 and 20 microm. The fractionated Er:YAG laser exhibited some of the same tissue interactions as its fully ablative counterparts. An increase in fluence resulted in an increase in ablative depth with minimal RTD. Additionally, RTD was unaffected by pulse stacking or by additional passes. Differences were that pulse stacking appeared to yield a more rapid decrease in ablation efficiency and additional passes did not seem to increase the depth of ablation.