Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Perspective
  • Published:

A “Gold Standard” Test for Diagnosing and Quantifying Hemolysis in Neonates and Infants

Journal of Perinatology volume 43, pages 1541–1547 (2023)Cite this article

Subjects

Abstract

Identifying “gold standard” diagnostic tests can promote evidence-based neonatology practice. Hemolysis is a pathological shortening of the erythrocyte lifespan, differing from erythrocyte senescence in responsible mechanisms and clinical implications. Diagnosing hemolysis goes beyond a binary (yes vs. no) determination. It is characterized according to magnitude, and as acute vs. chronic, and genetically based vs. not. For neonates with significant hyperbilirubinemia or anemia, detecting hemolysis and quantifying its magnitude provides diagnostic clarity. The 2022 American Academy of Pediatrics (AAP) Clinical Practice Guideline on management of hyperbilirubinemia in the newborn states that hemolysis is a risk factor for developing significant hyperbilirubinemia and neurotoxicity. The guideline recommends identifying hemolysis from any cause, but specific guidance is not provided. A spectrum of laboratory tests has been endorsed as diagnostic methods for hemolysis. Herein we examine these laboratory tests and recommend one as the “gold standard” for diagnosing and quantifying hemolysis in neonates and infants.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Fig. 1: Schematic representation of end-tidal breath carbon monoxide (ETCO) quantification for diagnosing and quantify hemolysis in a neonate or infant.

Similar content being viewed by others

References

  1. Bossuyt PM, Reitsma JB, Bruns DE, Gatsonis CA, Glasziou PP, Irwig LM, et al. Towards complete and accurate reporting of studies of diagnostic accuracy: the STARD initiative. Ann Intern Med. 2003;138:40–44.

    Article  PubMed  Google Scholar 

  2. Bossuyt PM, Reitsma JB, Bruns DE, Gatsonis CA, Glasziou PP, Irwig L, et al. STARD 2015: an updated list of essential items for reporting diagnostic accuracy studies. BMJ. 2015;351:h5527.

    Article  PubMed  PubMed Central  Google Scholar 

  3. Lantos JD. Ethical problems in decision making in the neonatal ICU. N Engl J Med. 2018;379:1851–60.

    Article  PubMed  Google Scholar 

  4. Lantos JD. The Belmont Report and innovative clinical research. Perspect Biol Med. 2020;63:389–400.

    Article  PubMed  Google Scholar 

  5. Means RT Jr, Glader BE. Anemia: general considerations. In: Greer JP, Arber DA, Appelbaum FR, Rogers GM, Glader B, List AF, et al. editors. Wintrobe’s clinical hematology. 14th ed. Philadelphia, PA: Wolters Kluwer; 2019. p. 603–12.

  6. Thiagarajan P, Parker CJ, Prchal JT. How do red blood cells die? Front Physiol. 2021;12:655393. https://doi.org/10.3389/fphys.2021.655393.

    Article  PubMed  PubMed Central  Google Scholar 

  7. Johnson L, Bhutani VK, Karp K, Sivieri EM, Shapiro SM. Clinical report from the pilot USA Kernicterus Registry (1992 to 2004). J Perinatol. 2009;29:S25–45.

    Article  PubMed  Google Scholar 

  8. Bhutani VK, Wong RJ. A global approach to prevent kernicterus: a dedication to Lois Johnson-Hamerman (Sep. 14, 1927, to Aug. 11, 2019). Semin Perinatol. 2021;45:151350.

    Article  PubMed  Google Scholar 

  9. Christensen RD, Agarwal AM, George TI, Bhutani VK, Yaish HM. Acute neonatal bilirubin encephalopathy in the State of Utah 2009-2018. Blood Cells Mol Dis. 2018;72:10–13.

    Article  PubMed  Google Scholar 

  10. Bhutani VK, Johnson-Hamerman L. The clinical syndrome of bilirubin-induced neurologic dysfunction. Semin Fetal Neonatal Med. 2015;20:6–13.

    Article  PubMed  Google Scholar 

  11. Christensen RD, Yaish HM. Hemolytic disorders causing severe neonatal hyperbilirubinemia. Clin Perinatol. 2015;42:515–27.

    Article  PubMed  Google Scholar 

  12. Christensen RD, Nussenzveig RH, Yaish HM, Henry E, Eggert LD, Agarwal AM. Causes of hemolysis in neonates with extreme hyperbilirubinemia. J Perinatol. 2014;34:616–9.

    Article  CAS  PubMed  Google Scholar 

  13. Christensen RD, Lambert DK, Henry E, Eggert LD, Yaish HM, Reading NS, et al. Unexplained extreme hyperbilirubinemia among neonates in a multihospital healthcare system. Blood Cells Mol Dis. 2013;50:105–9.

    Article  CAS  PubMed  Google Scholar 

  14. Bhutani VK, Stevenson DK. The need for technologies to prevent bilirubin-induced neurologic dysfunction syndrome. Semin Perinatol. 2011;35:97–100.

    Article  PubMed  Google Scholar 

  15. Maisels MJ, Newman TB. Kernicterus in otherwise health, breast-fed term newborns. Pediatrics. 1995;96:730–3.

    Article  CAS  PubMed  Google Scholar 

  16. Bhandari J, Thada PK, Yadav D. Crigler najjar syndrome. Treasure Island, FL: StatPearls Publishing; 2023.

  17. Strauss KA, Robinson DL, Vreman HJ, Puffenberger EG, Hart G, Morton DH. Management of hyperbilirubinemia and prevention of kernicterus in 20 patients with Crigler-Najjar disease. Eur J Pediatr. 2006;165:306–19.

    Article  PubMed  Google Scholar 

  18. Valsami S, Politou M, Boutsikou Τ, Briana D, Papatesta M, Malamitsi-Puchner A. Importance of direct antiglobulin test (DAT) in cord blood: causes of DAT (+) in a cohort study. Pediatr Neonatol. 2015;56:256–60.

    Article  PubMed  Google Scholar 

  19. Kaplan M, Hammerman C, Vreman HJ, Wong RJ, Stevenson DK. Direct antiglobulin titer strength and hyperbilirubinemia. Pediatrics. 2014;134:e1340–4.

    Article  PubMed  Google Scholar 

  20. Ross ME, Emery SP, de Alarcon PA, Watchko JF. Hemolytic disease of the fetus and newborn. maternal antibody mediated hemolysis. In: de Alarcon PA, Werner EJ, Christensen RD, Sola-Visner MC, editors. Neonatal hematology. Pathogenesis, diagnosis, and management of hematologic problems. 3rd ed. Cambridge: Cambridge University Press; 2021. p. 133–54.

  21. Moise KJ. Hemolytic disease of the fetus and newborn. In: Creasy RK, Resnik R, editors. Maternal-fetal medicine: principles and practice. 6th ed. Philadelphia, PA: WB Saunders; 2008. p. 477–503.

  22. Watchko JF. ABO hemolytic disease of the newborn: a need for clarity and consistency in diagnosis. J Perinatol. 2023;43:242–7.

    Article  CAS  PubMed  Google Scholar 

  23. Elsaie AL, Taleb M, Nicosia A, Zangaladze A, Pease ME, Newton K, et al. Comparison of end-tidal carbon monoxide measurements with direct antiglobulin tests in the management of neonatal hyperbilirubinemia. J Perinatol. 2020;40:1513–7.

    Article  CAS  PubMed  Google Scholar 

  24. Bahr TM, Shakib JH, Stipelman CH, Kawamoto K, Lauer S, Christensen RD. Improvement Initiative: end-tidal carbon monoxide measurement in newborns receiving phototherapy. J Pediatr. 2021;238:168–173.e2.

    Article  CAS  PubMed  Google Scholar 

  25. Christensen RD, Henry E, Bennett ST, Yaish HM. Reference intervals for reticulocyte parameters of infants during their first 90 days after birth. J Perinatol. 2016;36:61–6.

    Article  CAS  PubMed  Google Scholar 

  26. Hansen TWR, Wong RJ, Stevenson DK. Molecular physiology and pathophysiology of bilirubin handling by the blood, liver, intestine, and brain in the newborn. Physiol Rev. 2020;100:1291–346.

    Article  CAS  PubMed  Google Scholar 

  27. Bhutani VK, Wong RJ, Vreman HJ, Stevenson DK. Bilirubin production and hour-specific bilirubin levels. J Perinatol. 2015;35:735–8.

    Article  CAS  PubMed  Google Scholar 

  28. Petersen JR, Smith E, Okorodudu AO, Valbuena G, Bissell MG. Utilization of LDH isoenzymes in the diagnosis of myocardial infarction. Clin Lab Manage Rev. 1997;11:103–6.

    CAS  PubMed  Google Scholar 

  29. Yaish HM, Christensen RD, Lemons RS. Neonatal nonimmune hemolytic anemia. Curr Opin Pediatr. 2017;29:12–9.

    Article  PubMed  Google Scholar 

  30. Bahr TM, Judkins AJ, Christensen RD, Baer VL, Henry E, Minton SD, et al. Neonates with suspected microangiopathic disorders: performance of standard manual schistocyte enumeration the automated fragmented red cell count. J Perinatol. 2019;39:1555–61.

    Article  PubMed  Google Scholar 

  31. Bahr TM, Christensen TR, Henry E, Judkins AJ, Bennett ST, Pysher TJ, et al. Fragmented red blood cell counts of neonates with new-onset gastrointestinal disturbances. J Perinatol. 2022. https://doi.org/10.1038/s41372-022-01587-z.

  32. Christensen RD, Yaish HM, Lemons RS. Neonatal hemolytic jaundice: morphologic features of erythrocytes that will help you diagnose the underlying condition. Neonatology. 2014;105:243–9.

    Article  CAS  PubMed  Google Scholar 

  33. Gallagher PG. Diagnosis and management of rare congenital nonimmune hemolytic disease. Hematol Am Soc Hematol Educ Program. 2015;2015:392–9.

    Article  Google Scholar 

  34. Vreman HJ, Kwong LK, Stevenson DK. Carbon monoxide in blood: an improved microliter blood-sample collection system, with rapid analysis by gas chromatography. Clin Chem. 1984;30:1382–6.

    Article  CAS  PubMed  Google Scholar 

  35. Maisels MJ, Pathak A, Nelson NM, Nathan DG, Smith CA. Endogenous production of carbon monoxide in normal and erythroblastotic newborn infants. J Clin Invest. 1971;50:1–8.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Tidmarsh GF, Wong RJ, Stevenson DK. End-tidal carbon monoxide and hemolysis. J Perinatol. 2014;34:577–81.

    Article  CAS  PubMed  Google Scholar 

  37. Vreman HJ, Mahoney JJ, Stevenson DK. Carbon monoxide and carboxyhemoglobin. Adv Pediatr. 1995;42:303–4.

    CAS  PubMed  Google Scholar 

  38. Vreman HJ, Stevenson DK, Oh W, Fanaroff AA, Wright LL, Lemons JA, et al. Semiportable electrochemical instrument for determining carbon monoxide in breath. Clin Chem. 1994;40:1927–33.

    Article  CAS  PubMed  Google Scholar 

  39. Stevenson DK, Vreman HJ, Oh W, Fanaroff AA, Wright LL, Lemons JA, et al. Bilirubin production in healthy term infants as measured by carbon monoxide in breath. Clin Chem. 1994;40:1934–39.

    Article  CAS  PubMed  Google Scholar 

  40. Vreman HJ, Baxter LM, Stone RT, Stevenson DK. Evaluation of a fully automated end-tidal carbon monoxide instrument for breath analysis. Clin Chem. 1996;42:50–6.

    Article  CAS  PubMed  Google Scholar 

  41. Stevenson DK, Vreman HJ. Carbon monoxide and bilirubin production in neonates. Pediatrics. 1997;100:252–4.

    Article  CAS  PubMed  Google Scholar 

  42. Stevenson DK, Vreman HJ, Wong RJ, Contag CH. Carbon monoxide and bilirubin production in neonates. Semin Perinatol. 2001;25:85–93.

    Article  CAS  PubMed  Google Scholar 

  43. Vreman HJ, Wong RJ, Harmatz P, Fanaroff AA, Berman B, Stevenson DK. Validation of the Natus CO-Stat End Tidal Breath analyzer in children and adults. J Clin Monit Comput. 1999;15:421–7.

    Article  CAS  PubMed  Google Scholar 

  44. Krediet TG, Cirkel GA, Vreman HJ, Wong RJ, Stevenson DK, Groenendaal F, et al. End-tidal carbon monoxide measurements in infant respiratory distress syndrome. Acta Paediatr. 2006;95:1075–82.

    Article  PubMed  Google Scholar 

  45. Stevenson DK, Fanaroff AA, Maisels MJ, Young BW, Wong RJ, Vreman HJ, et al. Prediction of hyperbilirubinemia in near-term and term infants. Pediatrics. 2001;108:31–9.

    Article  CAS  PubMed  Google Scholar 

  46. American Academy of Pediatrics. Management of hyperbilirubinemia in the newborn infant 35 or more weeks of gestation. Pediatrics. 2004;114:297–16.

    Article  Google Scholar 

  47. James EB, Vreman HJ, Wong RJ, Stevenson DK, Vichinsky E, Schumacher L, et al. Elevated exhaled carbon monoxide concentration in hemoglobinopathies and its relation to red blood cell transfusion therapy. Pediatr Hematol Oncol. 2010;27:112–21.

    Article  CAS  PubMed  Google Scholar 

  48. Castillo Cuadrado ME, Bhutani VK, Aby JL, Vreman HJ, Wong RJ, Stevenson DK. Evaluation of a new end-tidal carbon monoxide monitor from the bench to the bedside. Acta Paediatr. 2015;104:e279–82.

    Article  CAS  PubMed  Google Scholar 

  49. Bhutani VK, Wong RJ, Vreman HJ, Stevenson DK, Jaundice Multinational Study Group. Bilirubin production and hour-specific bilirubin levels. J Perinatol. 2015;35:735–8.

    Article  CAS  PubMed  Google Scholar 

  50. Christensen RD, Lambert DK, Henry E, Yaish HM, Prchal JT. End-tidal carbon monoxide as an indicator of the hemolytic rate. Blood Cells Mol Dis. 2015;54:292–6.

    Article  CAS  PubMed  Google Scholar 

  51. Bhutani VK, Srinivas S, Castillo Cuadrado ME, Aby JL, Wong RJ, Stevenson DK. Identification of neonatal haemolysis: an approach to predischarge management of neonatal hyperbilirubinemia. Acta Paediatr. 2016;105:e189–94.

    Article  CAS  PubMed  Google Scholar 

  52. Christensen RD, Malleske DT, Lambert DK, Baer VL, Prchal JT, Denson LE, et al. Measuring end-tidal carbon monoxide of jaundiced neonates in the birth hospital to identify those with hemolysis. Neonatology. 2016;109:1–5.

    Article  CAS  PubMed  Google Scholar 

  53. Wong RJ, Bhutani VK, Stevenson DK. The importance of hemolysis and its clinical detection in neonates with hyperbilirubinemia. Curr Pediatr Rev. 2017;13:193–8.

    Article  CAS  PubMed  Google Scholar 

  54. Bhutani VK, Maisels MJ, Schutzman DL, Castillo Cuadrado ME, Aby JL, Bogen DL, et al. Identification of risk for neonatal haemolysis. Acta Paediatr. 2018;107:1350–6.

    Article  CAS  PubMed  Google Scholar 

  55. Bhatia A, Chua MC, Dela Puerta R, Rajadurai VS. Noninvasive detection of hemolysis with ETCOc measurement in neonates at risk for significant hyperbilirubinemia. Neonatology. 2020;117:612–8.

    Article  CAS  PubMed  Google Scholar 

  56. Du L, Ma X, Shen X, Bao Y, Chen L, Bhutani VK. Neonatal hyperbilirubinemia management: clinical assessment of bilirubin production. Semin Perinatol. 2021;45:151351.

    Article  PubMed  Google Scholar 

  57. Pakdeeto S, Christensen TR, Bahr TM, Gerday E, Sheffield MJ, Christensen KS, et al. Reference intervals for end-tidal carbon monoxide of preterm neonates. J Perinatol. 2022;42:116–20.

    Article  PubMed  Google Scholar 

  58. Bao Y, Zhu J, Ma L, Zhang H, Sun L, Xu C, et al. An end-tidal carbon monoxide nomogram for term and late-preterm chinese newborns. J Pediatr. 2022;250:16–21.e3.

    Article  PubMed  Google Scholar 

  59. Christensen RD, Bahr TM, Pakdeeto S, Supapannachart S, Zhang H. Perinatal hemolytic disorders, and identification using end tidal breath carbon monoxide. Curr Pediatr Rev. 2023;19:376–87.

    Article  PubMed  Google Scholar 

  60. Kemper AR, Newman TB, Slaughter JL, Maisels MJ, Watchko JF, Downs SM, et al. Clinical practice guideline revision: management of hyperbilirubinemia in the newborn infant 35 or more weeks of gestation. Pediatrics. 2022;150:e2022058859.

    Article  PubMed  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Division of Neonatology, Department of Pediatrics, University of Utah, Salt Lake City, UT, USA

    Robert D. Christensen & Timothy M. Bahr

  2. Obstetric and Neonatal Operations, Intermountain Health, Salt Lake City, UT, USA

    Robert D. Christensen & Timothy M. Bahr

  3. Department of Pediatrics, Division of Neonatal and Developmental Medicine, Stanford University School of Medicine, Stanford, CA, USA

    Ronald J. Wong, Hendrik J. Vreman, Vinod K. Bhutani & David K. Stevenson

Authors
  1. Robert D. Christensen
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Timothy M. Bahr
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Ronald J. Wong
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Hendrik J. Vreman
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Vinod K. Bhutani
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. David K. Stevenson
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

Conception and design: RDC and TMB. RDC wrote the initial draft of the manuscript. TMB, RJW, HJV, VKB, DKS reviewed all versions of the article and contributed to revising and improving the manuscript. All authors approved the final draft.

Corresponding author

Correspondence to Robert D. Christensen.

Ethics declarations

Competing interests

The authors declare no competing interests.

Additional information

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Christensen, R.D., Bahr, T.M., Wong, R.J. et al. A “Gold Standard” Test for Diagnosing and Quantifying Hemolysis in Neonates and Infants. J Perinatol 43, 1541–1547 (2023). https://doi.org/10.1038/s41372-023-01730-4

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/s41372-023-01730-4

This article is cited by

Search

Advanced search

Quick links