Storage lesions (SLs) occur when the red blood cell quality is altered during the preservation of blood units. Pre-storage leukoreduction would limit the number of SLs. The aims of this study were to evaluate the effectiveness of a leukoreduction filter for human use and the effect of pre-storage leukoreduction on some ematobiochemical parameters in stored canine whole blood. Seven canine blood units were tested. Each one was divided into two units—one leukoreduced (LRWB) and one non-leukoreduced (nLRWB). On each unit, we determined the complete blood count (CBC), lactate-dehydrogenase (LDH), electrolytes (Na+, K+, Cl−), morphological index (MI) and hemolysis, on storage days 0, 7, 14, 21, 28, 35, and 42. Leukoreduction allowed a 98.30% recovery of the RBC count, retaining 99.69% and 94.91% of WBCs and PLTs, respectively. We detected a significant increase of LDH and MI with strongly higher values in nLRWB compared to LRWB. A progressive increase in electrolytes and LDH concentrations was observed as indices of stored hemolysis. LDH showed significantly lower values in LRWB units compared to nLRWB, suggesting its release from leukocytes. In the majority of units, hemolysis reached 1% on the 42nd day of storage. We assert the human leukoreduction filter effectiveness on canine whole blood, and we recommend using nLRWB before day 14, especially for critically ill patients. The difference of the basal hemolysis (day 0) percentages observed between subjects suggests that more studies should be performed to confirm a possible inter-individual donor biological variability of RBC membrane resistance, as happens in humans.