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Determination of free heme in stored red blood cells with an apo-horseradish peroxidase-based assay

  • Vijith Vijayan , Robert Greite , Sebastian Schott , Julian Doricic , Kukuh Madyaningrana , Pooja Pradhan , Jörg Martens , Rainer Blasczyk , Sabina Janciauskiene and Stephan Immenschuh EMAIL logo
Published/Copyright: September 5, 2022
Biological Chemistry
From the journal Biological Chemistry Volume 403 Issue 11-12

Abstract

Transfusion effectiveness of red blood cells (RBCs) has been associated with duration of the storage period. Storage-dependent RBC alterations lead to hemolysis and release of toxic free heme, but the increase of free heme levels over time is largely unknown. In the current study, an apo-horseradish peroxidase (apoHRP)-based assay was applied to measure levels of free heme at regular intervals or periodically in supernatants of RBCs until a maximum storage period of 42 days. Free heme levels increased with linear time-dependent kinetics up to day 21 and accelerated disproportionally after day 28 until day 42, as determined with the apoHRP assay. Individual time courses of free heme in different RBC units exhibited high variability. Notably, levels of free hemoglobin, an established indicator of RBC damage, and those of total heme increased with continuous time-dependent linear kinetics over the entire 42 day storage period, respectively. Supernatants from RBC units with high levels of free heme led to inflammatory activation of human neutrophils. In conclusion, determining free heme in stored RBCs with the applied apoHRP assay may become feasible for testing of RBC storage quality in clinical transfusion medicine.

Keywords: apoHRP assay; free heme; red blood cells; storage lesion; transfusion
Corresponding author: Stephan Immenschuh, Institute of Transfusion Medicine and Transplant Engineering, Hannover Medical School, Carl-Neuberg-Str. 1, D-30625 Hannover, Germany, E-mail:

Funding source: Deutsche Forschungsgemeinschaft

Award Identifier / Grant number: IM 20/4-1

Acknowledgments

We wish to thank Anette Sarti for excellent technical support of the study.

  1. Author contributions: All the authors have accepted responsibility for the entire content of this submitted manuscript and approved submission.

  2. Research funding: Supported by grant IM 20/4-1 from the Deutsche Forschungsgemeinschaft, Bonn (Germany) [SI].

  3. Conflict of interest statement: The authors declare no conflicts of interest regarding this article.

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Supplementary Material

The online version of this article offers supplementary material (https://doi.org/10.1515/hsz-2022-0184).

Received: 2022-05-17
Revised: 2022-08-10
Accepted: 2022-08-15
Published Online: 2022-09-05
Published in Print: 2022-11-25

© 2022 Walter de Gruyter GmbH, Berlin/Boston

Articles in the same Issue

  1. Frontmatter
  2. Heme research – the past, the present and the future
  3. A primer on heme biosynthesis
  4. New roles for GAPDH, Hsp90, and NO in regulating heme allocation and hemeprotein function in mammals
  5. The role of host heme in bacterial infection
  6. Signal transduction mechanisms in heme-based globin-coupled oxygen sensors with a focus on a histidine kinase (AfGcHK) and a diguanylate cyclase (YddV or EcDosC)
  7. Heme delivery to heme oxygenase-2 involves glyceraldehyde-3-phosphate dehydrogenase
  8. Novel insights into heme binding to hemoglobin
  9. Extracellular hemin is a reverse use-dependent gating modifier of cardiac voltage-gated Na+ channels
  10. Assessment of the breadth of binding promiscuity of heme towards human proteins
  11. Determination of free heme in stored red blood cells with an apo-horseradish peroxidase-based assay
  12. Comparative studies of soluble and immobilized Fe(III) heme-peptide complexes as alternative heterogeneous biocatalysts
https://doi.org/10.1515/hsz-2022-0184
Keywords for this article
apoHRP assay; free heme; red blood cells; storage lesion; transfusion

Articles in the same Issue

  1. Frontmatter
  2. Heme research – the past, the present and the future
  3. A primer on heme biosynthesis
  4. New roles for GAPDH, Hsp90, and NO in regulating heme allocation and hemeprotein function in mammals
  5. The role of host heme in bacterial infection
  6. Signal transduction mechanisms in heme-based globin-coupled oxygen sensors with a focus on a histidine kinase (AfGcHK) and a diguanylate cyclase (YddV or EcDosC)
  7. Heme delivery to heme oxygenase-2 involves glyceraldehyde-3-phosphate dehydrogenase
  8. Novel insights into heme binding to hemoglobin
  9. Extracellular hemin is a reverse use-dependent gating modifier of cardiac voltage-gated Na+ channels
  10. Assessment of the breadth of binding promiscuity of heme towards human proteins
  11. Determination of free heme in stored red blood cells with an apo-horseradish peroxidase-based assay
  12. Comparative studies of soluble and immobilized Fe(III) heme-peptide complexes as alternative heterogeneous biocatalysts
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