The objective of this study was to investigate the possible existence of a set of critical combinations among medical CO2 laser device set-ups and to compare the resigns of diameter and depth of irradiated tissue. These data are useful to the surgeon to identify a general operative protocol that allows both to forecast the beam behavior on a large variety of operative conditions accurately and to identify the related safety margins. Two methods have been addressed: the more traditional free-air beam delivery to the tissue and the more modern requirements of the minimally invasive surgery concepts, which use fiberoptic-based delivery systems for the CO2 laser beam. In the past many articles have been written about the interaction between laser beams and biological media. However, the CO2 laser beam at 10.6 microm has always challenged research institutions and manufacturers to find the ideal combinations between a suitable fiberoptic-based delivery system concept and the injury threshold conditions recommended for minimally invasive surgery (MIS) and transmyocardial laser revascularization (TMLR) applications. One of the main challenges is to deliver the radiation at 10.6 microm without serious damage to the fiberoptic-based scalpel and the irradiated tissue by the burn of the fiber in use. Five rabbits weighing 3 to 3.5 kg were sacrificed and 60 samples of trachea, myocardium, aorta, and esophagus were immediately excised, trimmed free of the adherent connective tissue, and irradiated in the intima portion of the wall. A commercial TEM medical CO2 laser was coupled to a silver halide fiber (0.9 mm in diameter) and subsequently to regular focusing heads (2.5-inch and 5-inch focal length) for pulsed laser beam delivery. The parameters chosen were: 33, 50, 65, and 100 mJ per pulse, 10 and 20 ms pulse width, delivered in two sets of experiments, one with all the pulsed beam frequencies below 5 Hz, the other between 15 Hz and 20 Hz. The same tests were conducted on 10 blocks of 3 different plastic types to simulate in one single procedure the laser radiation in responses to both hard, low-water content tissues such as bone and of common plastic compounds (such as polymethylmethacrylates [PMMA]) routinely used in orthopedic surgery. An optical microscope was used to measure all the lesions (diameter and depth in millimeters) in all the samples and to identify the smallest and the largest one against which similar thermal injuries found on the other media were compared. This study demonstrates that for power densities between about 520 and 790 W/cm2 per pulse achieved with the silver halide fiber generated a minimal lesion on the myocardium and aortic tissues. This can be used as reference threshold for MIS and TMLR. The minimal injury threshold on PMMA has been reached at 393 W/cm2 per pulse at 1 Hz. This approached the conditions of a pulsed beam delivered in air on the same medium via a focal spot of an 8.7 inch focal. Surprisingly, all the treated media show lesions that follow similar patterns within a well-defined and limited range of both diameter and depth. This effect was obtained by using power densities ranging from 393 to 6310 W/cm2 per pulse regardless of all the other parameters, including power delivery method, the type of irradiated tissue and the frequency between 0 and 20 Hz. Only the combination power density-type of tissue appears to be decisive. The geometrical convergence of the diameter shows a much smoother pattern than the one of the depth, due to two different irradiation modalities. The selection of the CO2 laser beam parameters and the irradiated media reported in this article have allowed identification of a critical set of ablative conditions to be further used in the operating room.