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Towards the development of a certified reference material for hemoglobin A2

  • Renata Paleari , Amalia Muñoz , Andrea Mosca and on behalf of the IFCC Working Group on Standardization of HbA2 (WG-HbA2)
Published/Copyright: October 30, 2010
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Clinical Chemistry and Laboratory Medicine
From the journal Clinical Chemistry and Laboratory Medicine Volume 48 Issue 11

Abstract

Background: In 2004, a working group on the standardization of hemoglobin A2 (HbA2) was created within the IFCC, with the aim of developing a reference system for this analyte. One goal was to prepare a certified reference material in collaboration with the Institute for Reference Materials and Measurements (IRMM). This paper describes the properties of a first batch of this candidate study material.

Methods: Eighty millilitre of fresh whole blood, collected from a healthy blood donor, was treated by removing plasma, white blood cells and platelets. Red cells were hemolyzed to prepare 100 vials of lyophilized material (approximately 155 mg per vial). After reconstitution, the HbA2 content was measured with a total of seven HPLC methods, three electrophoretic techniques, and two capillary electrophoresis (CE) methods. Homogeneity was tested in a subset of five vials. Stability during storage at +4°C and –20°C was tested monthly over a period of 1 year. The commutability of this material was assessed by analysing the study material together with a set of 54 fresh blood samples, with a subgroup of the above mentioned methods, only by one routine HPLC (Bio-Rad Variant II, dual kit) and by a CE method (Beckman PA800, Analis kit), respectively.

Results: The chromatographic and electrophoretic patterns obtained by all the HPLC, electrophoretic and CE techniques did not show any difference between those obtained using the first study material and those obtained with fresh blood samples. The lot was found to be homogeneous on the basis of the content of lyophilized powder per vial. The HbA2 concentration in the lyophilized material remained stable at +4°C and –20°C, even after 1 year of storage. After reconstitution, the HbA2 concentration did not change for more than 2 weeks in the refrigerator at +4°C. The normalized residual of the study material, measuring the degree of its commutability was 0.9, similar to that obtained on other home prepared and some commercial controls.

Conclusions: Ideally, fresh whole blood is the best reference material in the meterological traceability chain for HbA2 analysis. However, for a number of reasons the preparation of large batches of fresh whole blood to be used as secondary reference material for HbA2 is not practical. In our work, we have proven that lyophilization does not appear to cause any matrix effect or inhomogeneity in the study material, which also confirmed to be commutable for the Bio-Rad Variant II (dual kit) and Beckman PA800 (Analis kit) methods. We conclude that a material similarly prepared as the current study material and value assigned with the candidate reference measurement procedure still under development will be suitable to calibrate various routine methods for HbA2. This will result in improvement of the inter-method variability for this important biochemical marker.

Clin Chem Lab Med 2010;48:1611–8.

Keywords: commutability; HbA2; reference materials; standardization; thalassemia
Corresponding author: Prof. Andrea Mosca, Dip. di Scienze e Tecnologie Biomediche, Via Fratelli Cervi 93, 20090 Segrate, Milan, Italy Phone: +39 02 5033 0422, Fax +39 02 9998 7559,
Published Online: 2010-10-30
Published in Print: 2010-11-01

©2010 by Walter de Gruyter Berlin New York

Articles in the same Issue

  1. Editorial
  2. IFCC Scientific Division
  3. IFCC Documents
  4. Nucleic acid reference materials (NARMs): definitions and issues
  5. An appraisal of statistical procedures used in derivation of reference intervals
  6. An outline for a vocabulary of nominal properties and examinations – basic and general concepts and associated terms
  7. Review
  8. Standardizing plasma protein measurements worldwide: a challenging enterprise
  9. Status Reports
  10. Standardization activities in the field of thyroid function tests: a status report
  11. Toward standardization of carbohydrate-deficient transferrin (CDT) measurements: II. Performance of a laboratory network running the HPLC candidate reference measurement procedure and evaluation of a candidate reference material
  12. Research Articles
  13. Common reference intervals for aspartate aminotransferase (AST), alanine aminotransferase (ALT) and γ-glutamyl transferase (GGT) in serum: results from an IFCC multicenter study
  14. Development of a candidate secondary reference procedure (immunoassay based measurement procedure of higher metrological order) for cardiac troponin I: I. Antibody characterization and preliminary validation
  15. Towards the development of a certified reference material for hemoglobin A2
  16. Short Communication
  17. First certified reference material for cystatin C in human serum ERM-DA471/IFCC
  18. General Clinical Chemistry and Laboratory Medicine
  19. Increased γ-glutamyltransferase and decreased total bilirubin are associated with metabolic syndrome in Korean postmenopausal women
  20. Effect of homeostasis model assessment computational method on the definition and associations of insulin resistance
  21. Association between thyroid hormones, lipids and oxidative stress biomarkers in overt hypothyroidism
  22. Biochemical markers of particle induced osteolysis in C57BL/6 mice
  23. An on-line solid phase extraction procedure for the routine quantification of urinary methylmalonic acid by liquid chromatography-tandem mass spectrometry
  24. Cancer Diagnostics
  25. Increased integrity of circulating cell-free DNA in plasma of patients with acute leukemia
  26. The relationship between three heat shock protein 70 gene polymorphisms and susceptibility to lung cancer
  27. Real-time polymerase chain reaction quantification of free DNA in serum of patients with polyps and colorectal cancers
  28. Letters to the Editor
  29. Thyroid hormones are stable even during prolonged frozen storage
  30. Thyroid hormones after long-term cold storage: stable or unstable, that is the question
  31. Biomarkers for early detection of mesothelioma in asbestos-exposed subjects
  32. Mean platelet volume and platelet distribution width are useful in the differential diagnosis of aplastic anemia and idiopathic thrombocytopenic purpura
  33. Kinetics of highly sensitive troponin I and T after eccentric exercise
  34. Interferences in red blood cell counting in urinalysis using evacuated tubes
  35. Congress Abstracts
  36. 3rd Annual Congress of the Austrian Society for Laboratory Medicine and Clinical Chemistry Salzburg, Austria, 27-30 October 2010
https://doi.org/10.1515/CCLM.2010.317
Keywords for this article
commutability; HbA2; reference materials; standardization; thalassemia

Articles in the same Issue

  1. Editorial
  2. IFCC Scientific Division
  3. IFCC Documents
  4. Nucleic acid reference materials (NARMs): definitions and issues
  5. An appraisal of statistical procedures used in derivation of reference intervals
  6. An outline for a vocabulary of nominal properties and examinations – basic and general concepts and associated terms
  7. Review
  8. Standardizing plasma protein measurements worldwide: a challenging enterprise
  9. Status Reports
  10. Standardization activities in the field of thyroid function tests: a status report
  11. Toward standardization of carbohydrate-deficient transferrin (CDT) measurements: II. Performance of a laboratory network running the HPLC candidate reference measurement procedure and evaluation of a candidate reference material
  12. Research Articles
  13. Common reference intervals for aspartate aminotransferase (AST), alanine aminotransferase (ALT) and γ-glutamyl transferase (GGT) in serum: results from an IFCC multicenter study
  14. Development of a candidate secondary reference procedure (immunoassay based measurement procedure of higher metrological order) for cardiac troponin I: I. Antibody characterization and preliminary validation
  15. Towards the development of a certified reference material for hemoglobin A2
  16. Short Communication
  17. First certified reference material for cystatin C in human serum ERM-DA471/IFCC
  18. General Clinical Chemistry and Laboratory Medicine
  19. Increased γ-glutamyltransferase and decreased total bilirubin are associated with metabolic syndrome in Korean postmenopausal women
  20. Effect of homeostasis model assessment computational method on the definition and associations of insulin resistance
  21. Association between thyroid hormones, lipids and oxidative stress biomarkers in overt hypothyroidism
  22. Biochemical markers of particle induced osteolysis in C57BL/6 mice
  23. An on-line solid phase extraction procedure for the routine quantification of urinary methylmalonic acid by liquid chromatography-tandem mass spectrometry
  24. Cancer Diagnostics
  25. Increased integrity of circulating cell-free DNA in plasma of patients with acute leukemia
  26. The relationship between three heat shock protein 70 gene polymorphisms and susceptibility to lung cancer
  27. Real-time polymerase chain reaction quantification of free DNA in serum of patients with polyps and colorectal cancers
  28. Letters to the Editor
  29. Thyroid hormones are stable even during prolonged frozen storage
  30. Thyroid hormones after long-term cold storage: stable or unstable, that is the question
  31. Biomarkers for early detection of mesothelioma in asbestos-exposed subjects
  32. Mean platelet volume and platelet distribution width are useful in the differential diagnosis of aplastic anemia and idiopathic thrombocytopenic purpura
  33. Kinetics of highly sensitive troponin I and T after eccentric exercise
  34. Interferences in red blood cell counting in urinalysis using evacuated tubes
  35. Congress Abstracts
  36. 3rd Annual Congress of the Austrian Society for Laboratory Medicine and Clinical Chemistry Salzburg, Austria, 27-30 October 2010
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