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Introduction: Hemolysis in the emergency department (ED) can significantly delay results and appropriate action. We evaluated the main sources of hemolysis during sample collection, and to evaluate the use of rapid serum tubes (RST) as a transport hemolysis-mitigating measure for high-sensitivity troponin T (hs-cTnT) testing., Methods: We examined the effect of tube type, tube fill, types of sample draw and collection methods on hemolysis and hs-cTnT in samples (n = 158) from ED patients. We also compared hs-cTnT values in paired RST and plasma separate tube (PST) samples that were hemolysis-free., Results: The primary source of hemolysis in samples collected in the ED was underfilling tubes. In both tube types, PST and RST, filled tubes showed a median reduction in hemolysis of 69.1 % (p < 0.0001). Blood collected in RST also experienced less hemolysis compared to PST. In hemolysis-free samples, false positive results in PST were noted in patients with hs-cTnT values < 50 ng/l., Conclusion: We suggest that proper tube filling during sample collection and use of RST tubes can significantly reduce the effects of hemolysis. In addition, laboratories should be aware that PST tubes have a non-trivial rate of false positives when hs-cTnT < 50 ng/l., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)