Little is known about the potential consequences of pancreatic tissue sampling in dogs. The goal of the present study was to evaluate changes in serum trypsin-like immunoreactivity and canine-specific pancreatic lipase after pancreatic fine-needle aspiration and surgical biopsy in 27 clinically healthy dogs. Presurgical, ultrasound-guided aspiration of the pancreas was performed with the dogs under sedation. Subsequently, all the dogs underwent intraoperative pancreatic fine-needle aspiration and clamshell biopsy. After euthanasia, pancreata were sectioned for histopathologic evaluation. Serum pancreatic enzyme levels were measured at 3 time points: baseline, after ultrasound-guided aspiration, and after intraoperative aspiration and biopsy. No significant differences were detected among mean serum pancreatic lipase values at any point (P > 0.05). Serum trypsin-like immunoreactivity did not change from baseline (18.2 +/- 2.1 microg/dl; mean +/- standard error) after ultrasound-guided aspiration (13.6 +/- 2.2 microg/dl) but increased significantly after intraoperative sampling (44.8 +/- 1.9 microg/dl; P < 0.0005). After surgical biopsy, the 20 dogs that had both ultrasound-guided and intraoperative sampling had a higher mean (SE) serum trypsin-like immunoreactivity (44.8 +/- 1.9 microg/dl) than the 7 dogs that had only intraoperative samples taken (36.4 +/- 4.1 microg/dl; P < 0.05). All 27 pancreata were grossly normal before intraoperative sampling. Pancreatic sampling was associated with increased serum trypsin-like immunoreactivity and mild, peracute necrosis, inflammation, hemorrhage, and fibrin deposition. Tissue damage from sampling was not sufficient to cause an elevation in canine-specific pancreatic lipase in the time frame evaluated. Further studies are needed to determine longer-term effects of pancreatic sampling on enzyme levels and clinical outcome.