When Hemolysis Misleads: Implications for the Clinical Interpretation of Enzyme Activities in Pediatric Samples

Background and Objectives: Hemolysis is one of the most frequent preanalytical interferences in clinical laboratory practice. This laboratory-based experimental study evaluated the effect of different degrees of hemolysis on plasma enzyme activities in pediatric blood samples. Materials and Methods: Twenty-six venous blood samples collected in lithium-heparin tubes were gently homogenized and divided into four equal whole-blood aliquots. One aliquot served as the untreated reference, while the remaining three aliquots were subjected to two, four, or eight aspiration-reinjection cycles through an 18-gauge needle to induce increasing degrees of erythrocyte lysis. All four aliquots were subsequently centrifuged under identical conditions and classified according to the semiquantitative hemolysis component of the analyzer-generated LIH index as non-hemolyzed, H1, H2, H3, H4, or H5. Activities of alpha-amylase, alkaline phosphatase, alanine aminotransferase, aspartate aminotransferase, gamma-glutamyl transferase, creatine kinase, and lactate dehydrogenase, as well as potassium concentration, were measured using the same DxC 700 AU automated clinical chemistry analyzer. Statistical analysis included 26 samples and was performed using the Wilcoxon signed-rank test. Results: Hemolysis caused a progressive increase in lactate dehydrogenase, aspartate aminotransferase, creatine kinase, alanine aminotransferase, and potassium, with the strongest effects observed for lactate dehydrogenase and aspartate aminotransferase. Alpha-amylase and alkaline phosphatase showed a decreasing trend, while gamma-glutamyl transferase showed variable behavior across hemolysis groups. Conclusions: The findings confirm that even low degrees of hemolysis can substantially alter selected biochemical results, especially lactate dehydrogenase, aspartate aminotransferase, and creatine kinase. Laboratory reports from hemolyzed pediatric samples should therefore be interpreted with caution to reduce the risk of clinically misleading results.