Objective' To determine whether percentages of neutrophils in tracheal aspirate (TA) samples collected from racehorses are increased after exercise and whether interpretation of results from TA samples taken before and after exercise agree.Design' Case series of 40 young Thoroughbred and Standardbred racehorses in race training presented for evaluation of poor performance.Procedure' TA samples were collected endoscopically from racehorses presented for poor performance 24 h before and 1 to 2 h after high speed treadmill exercise testing. Aliquots of the retrieved fluid were cytocentrifuged and smears were stained with Diff-QuikÂ®. Mean neutrophil counts were expressed as percentages of the total number of inflammatory cells counted and subsequently were categorised as either above or below an accepted cut-off of 20%. Comparisons between percentages of neutrophils before and after exercise were made.Results' Percentage of neutrophils from TA samples obtained from racehorses after exercise was significantly higher than neutrophil percentages from TA samples collected from the same horse before exercise. In horses with TA specimens that were categorised as having ' 20% neutrophils before treadmill exercise, the percentage of neutrophils in their TA specimens after exercise was, on average, significantly higher and was greater than the cut-off value of 20%.Conclusion' Recent strenuous exercise may change the proportion of neutrophils in lower airways of racehorses and practitioners should be aware of this when collecting and interpreting the results from TA samples. The most practical time for collection of a TA sample to obtain the most diagnostically useful information might be after a suitable washout period of at least 1 to 2 h post-exercise.