Startseite Medizin A comparative evaluation of the analytical performances of premier resolution-high-performance liquid chromatography (PR-HPLC) with capillary zone electrophoresis (CZE) assays for the detection of hemoglobin variants and the quantitation of HbA0, A2, E, and F
Artikel
Lizenziert
Nicht lizenziert Erfordert eine Authentifizierung

A comparative evaluation of the analytical performances of premier resolution-high-performance liquid chromatography (PR-HPLC) with capillary zone electrophoresis (CZE) assays for the detection of hemoglobin variants and the quantitation of HbA0, A2, E, and F

  • Sirikwan Laksap , Suphisara Suanboon , Manoo Punyamung , Chedtapak Ruengdit und Sakorn Pornprasert EMAIL logo
Veröffentlicht/Copyright: 11. Januar 2024
Clinical Chemistry and Laboratory Medicine (CCLM)
Aus der Zeitschrift Clinical Chemistry and Laboratory Medicine (CCLM) Band 62 Heft 7

Abstract

Objectives

Hemoglobinopathies, including thalassemia and hemoglobin (Hb) variants, are common hematological disorders in tropical countries. Accurate and precise separation of hemoglobin types and reliable quantitation are necessary for differential diagnosis of these disorders.

Methods

We have evaluated the analytical performances of premier resolution-high-performance liquid chromatography (PR-HPLC; Trinity Biotech, Co. Wicklow, Ireland) to assist in the presumptive diagnosis of thalassemia and Hb variants commonly found in Southeast Asian countries. HbA0, HbA2, HbE, and HbF levels were separated and quantified in 120 blood samples from unrelated adult subjects and compared with those analyzed by capillary zone electrophoresis (CZE; CAPILLARYS™ 2, Sebia, Norcross, GA, US). The Hb analysis patterns of Hb variants obtained from the PR-HPLC system were also compared to those obtained from HPLC (VARIANT II, β-thalassemia Short Program, Bio-Rad, Laboratories, Hercules, CA, US) and CZE systems.

Results

The PR-HPLC had excellent precision with a coefficient of variation (CV) for HbA2 quantitation of 3.8 % within-run and 5.2 % between-run. The levels of HbA2/E quantified by the PR-HPLC system correlated well with those of the CZE system (r=0.997). In addition, thalassemia interpretation results obtained from the PR-HPLC and the CZE showed 100 % agreement. Moreover, chromatograms of the PR-HPLC were also comparable to those of VII-HPLC and CAP2-CZE electropherograms.

Conclusions

The PR-HPLC system would be applicable to diagnose common forms of thalassemia and Hb variants in Southeast Asia.

Keywords: capillary zone electrophoresis; hemoglobin analysis; hemoglobinopathy; high-performance liquid chromatography; premier resolution; thalassemia
Corresponding author: Dr. Sakorn Pornprasert, Department of Medical Technology, Faculty of Associated Medical Sciences, Chiang Mai University, 110 Intawaroros Road, Chiang Mai, 50200, Thailand, Phone: +66 53 945078, Fax: +66 53 946042, E-mail:

Acknowledgments

The authors thank technicians in the Associated Medical Sciences Clinical Service Center, Faculty of Associated Medical Sciences, Chiang Mai University, Chiang Mai, Thailand, for their help and assistance. We also would like to thank Biozen Co., Ltd. and staffs for supplying reagents and automatic analyzers.

  1. Research ethics: This study was submitted for Ethical Approval from the Ethics Committee of the Faculty of Associated Medical Sciences, Chiang Mai University, Thailand (AMSEC-66EM-025).

  2. Informed consent: The Ethics Committee waived the need for consent as samples were leftover specimens from the routine, and the samples and associated data could not be traced back to the subjects. In addition, information that identified individual subjects could not be accessed during or after data collection.

  3. Author contributions: Sakorn Pornprasert and Chedtapak Ruengdit designed the study. Sirikwan Laksap, Suphisara Suanboon, Manoo Punyamung, and Chedtapak Ruengdit performed experiments and analyzed data. Sakorn Pornprasert, Sirikwan Laksap, and Suphisara Suanboon discussed the study and wrote the manuscript. The authors have accepted responsibility for the entire content of this manuscript and approved its submission.

  4. Competing interests: The authors state no conflicts of interest. The authors alone are responsible for the content and writing of the paper.

  5. Research funding: None declared. This research received no specific grant from public, commercial, or not-for-profit funding agencies.

  6. Data availability: Not applicable.

References

1. Fucharoen, S, Winichagoon, P. Thalassemia and abnormal hemoglobin. Int J Hematol 2002;76(2 Suppl):83–9. https://doi.org/10.1007/bf03165094.Suche in Google Scholar

2. Fucharoen, S, Winichagoon, P. New updating into hemoglobinopathies. Int J Lab Hematol 2012;34:559–65. https://doi.org/10.1111/j.1751-553x.2012.01446.x.Suche in Google Scholar

3. Cao, A, Galanello, R, Rosatelli, MC. Prenatal diagnosis and screening of the haemoglobinopathies. Baillieres Clin Haematol 1998;11:215–38. https://doi.org/10.1016/s0950-3536(98)80076-0.Suche in Google Scholar PubMed

4. Chan, LC, Ma, SK, Chan, AY, Ha, SY, Waye, JS, Lau, YL, et al.. Should we screen for globin gene mutations in blood samples with mean corpuscular volume (MCV) greater than 80 fL in areas with a high prevalence of thalassaemia? J Clin Pathol 2001;54:317–20. https://doi.org/10.1136/jcp.54.4.317.Suche in Google Scholar PubMed PubMed Central

5. Winichagoon, P, Thitivichianlert, A, Lebnak, T, Piankijagum, A, Fucharoen, S. Screening for the carriers of thalassemias and abnormal hemoglobins at the community level. Southeast Asian J Trop Med Publ Health 2002;33:145–50.Suche in Google Scholar

6. Fucharoen, S, Winichagoon, P, Wisedpanichkij, R, Sae-Ngow, B, Sriphanich, R, Oncoung, W, et al.. Prenatal and postnatal diagnoses of thalassemias and hemoglobinopathies by HPLC. Clin Chem 1998;44:740–8. https://doi.org/10.1093/clinchem/44.4.740.Suche in Google Scholar

7. Winichagoon, P, Svasti, S, Munkongdee, T, Chaiya, W, Boonmongkol, P, Chantrakul, N, et al.. Rapid diagnosis of thalassemias and other hemoglobinopathies by capillary electrophoresis system. Transl Res 2008;152:178–84. https://doi.org/10.1016/j.trsl.2008.08.004.Suche in Google Scholar PubMed

8. Van Delft, P, Lenters, E, Bakker-Verweij, M, de Korte, M, Baylan, U, Harteveld, CL, et al.. Evaluating five dedicated automatic devices for haemoglobinopathy diagnostics in multi-ethnic populations. Int J Lab Hematol 2009;31:484–95. https://doi.org/10.1111/j.1751-553x.2009.01158.x.Suche in Google Scholar PubMed

9. Liao, C, Zhou, JY, Xie, XM, Li, J, Li, R, Li, DZ. Detection of hb constant spring by a capillary electrophoresis method. Hemoglobin 2010;34:175–8. https://doi.org/10.3109/03630261003680191.Suche in Google Scholar PubMed

10. Waneesorn, J, Panyasai, S, Kongthai, K, Singboottra, P, Pornprasert, S. Comparison between capillary electrophoresis and high performance liquid chromatography for detection and quantification of Hb Constant Spring [HbCS; α142, Term→Gln (TAA>CAA IN α2)]. Hemoglobin 2011;35:338–45. https://doi.org/10.3109/03630269.2011.588140.Suche in Google Scholar PubMed

11. Satthakarn, S, Panyasai, K, Phasit, A, Panyasai, S. Reliability of hemoglobin A(2) value as measured by the Premier Resolution system for screening of beta-thalassemia carriers. Clin Chem Lab Med 2024;62:453–63. https://doi.org/10.1515/cclm-2023-1006.Suche in Google Scholar PubMed

12. Achour, A, de Grouw, E, van Erp, F, Arkesteijn, S, Schaap, R, Huurne, JT, et al.. The first report of hemoglobin E in combination with the highly unstable alpha-globin variant Hb Adana: the importance of molecular confirmation. Int J Lab Hematol 2019;41:e76–8. https://doi.org/10.1111/ijlh.12965.Suche in Google Scholar PubMed PubMed Central

13. Ruengdit, C, Punyamung, M, Intasai, N, Pornprasert, S. Single-tube multiplex real-time PCR with EvaGreen and high-resolution melting analysis for diagnosis of alpha0-thalassemia--SEA,--THAI, and--CR type deletions. PLoS One 2023;18:e0293838. https://doi.org/10.1371/journal.pone.0293838.Suche in Google Scholar PubMed PubMed Central

14. The Ministry of Public Health. Guidelines for laboratory testing to support the control and prevention of thalassemia. Nonthaburi: Department of Medical Sciences, Ministry of Public Health; 2019.Suche in Google Scholar

15. Pornprasert, S, Kongthai, K, Waneesorn, J, Jaiping, K, Treesuwan, K. Development of control material for hemoglobin analysis. Clin Chem Lab Med 2011;50:351–6. https://doi.org/10.1515/cclm.2011.777.Suche in Google Scholar PubMed

16. Tangvarasittichai, S, Tangvarasittichai, O, Jermnim, N. Comparison of fast protein liquid chromatography (FPLC) with HPLC, electrophoresis & microcolumn chromatography techniques for the diagnosis of beta-thalassaemia. Indian J Med Res 2009;129:242–8.Suche in Google Scholar

17. Sangkitporn, S, Sangkitporn, SK, Tanjatham, S, Suwannakan, B, Rithapirom, S, Yodtup, C, et al.. Multicenter validation of fully automated capillary electrophoresis method for diagnosis of thalassemias and hemoglobinopathies in Thailand. Southeast Asian J Trop Med Publ Health 2011;42:1224–32.Suche in Google Scholar

18. Munkongdee, T, Chen, P, Winichagoon, P, Fucharoen, S, Paiboonsukwong, K. Update in laboratory diagnosis of thalassemia. Front Mol Biosci 2020;7:74. https://doi.org/10.3389/fmolb.2020.00074.Suche in Google Scholar PubMed PubMed Central

19. Jindadamrongwech, S, Tungbuppha, N, Chuncharunee, S, Butthep, P. Hb Tak and HbQ-Thailand in Thai patients are S-window hemoglobin variants revealed by high performance liquid chromatography. Hemoglobin 2010;34:161–4. https://doi.org/10.3109/03630261003679631.Suche in Google Scholar PubMed

20. Pornprasert, S, Punyamung, M. Detection of compound heterozygous of hb constant spring and hb q-Thailand by capillary electrophoresis and high performance liquid chromatography. Indian J Hematol Blood Transfus 2015;31:229–32. https://doi.org/10.1007/s12288-014-0411-3.Suche in Google Scholar PubMed PubMed Central

21. Riou, J, Szuberski, J, Godart, C, Wajcman, H, Oliveira, JL, Hoyer, JD, et al.. Precision of CAPILLARYS 2 for the detection of hemoglobin variants based on their migration positions. Am J Clin Pathol 2018;149:172–80. https://doi.org/10.1093/ajcp/aqx148.Suche in Google Scholar PubMed

22. Srivorakun, H, Singha, K, Fucharoen, G, Sanchaisuriya, K, Fucharoen, S. A large cohort of hemoglobin variants in Thailand: molecular epidemiological study and diagnostic consideration. PLoS One 2014;9:e108365. https://doi.org/10.1371/journal.pone.0108365.Suche in Google Scholar PubMed PubMed Central

23. Keren, DF, Hedstrom, D, Gulbranson, R, Ou, CN, Bak, R. Comparison of Sebia Capillarys capillary electrophoresis with the Primus high-pressure liquid chromatography in the evaluation of hemoglobinopathies. Am J Clin Pathol 2008;130:824–31. https://doi.org/10.1309/ajcpqy80hzwhhgzf.Suche in Google Scholar PubMed

24. Pornprasert, S, Treesuwan, K, Punyamung, M, Kongthai, K. HbA2/E levels found in co-inheritance with the α-thalassemia-1 - -(SEA)/type deletion and either HbE or β-thalassemia. Hemoglobin 2012;36:381–7. https://doi.org/10.3109/03630269.2012.679375.Suche in Google Scholar PubMed

25. Pornprasert, S, Jaiping, K. Blood transfusion from a HbE trait donor can affect beta-thalassemia diagnosis. Hemoglobin 2014;38:295–8. https://doi.org/10.3109/03630269.2014.926913.Suche in Google Scholar PubMed

Received: 2023-12-17
Accepted: 2024-01-04
Published Online: 2024-01-11
Published in Print: 2024-06-25

© 2024 Walter de Gruyter GmbH, Berlin/Boston

Artikel in diesem Heft

  1. Frontmatter
  2. Editorial
  3. LC-MS/MS random access automation – a game changer for the 24/7 clinical laboratory
  4. Reviews
  5. Neurofilament light protein as a biomarker for spinal muscular atrophy: a review and reference ranges
  6. Differential diagnosis of ascites: etiologies, ascitic fluid analysis, diagnostic algorithm
  7. Opinion Papers
  8. Clinical Decision Support System in laboratory medicine
  9. Blood over-testing: impact, ethical issues and mitigating actions
  10. General Clinical Chemistry and Laboratory Medicine
  11. An isotope dilution-liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS)-based candidate reference measurement procedure for the quantification of zonisamide in human serum and plasma
  12. An isotope dilution-liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS)-based candidate reference measurement procedure for the quantification of carbamazepine in human serum and plasma
  13. An isotope dilution-liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS)-based candidate reference measurement procedure (RMP) for the quantification of phenobarbital in human serum and plasma
  14. An isotope dilution-liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS)-based candidate reference measurement procedure (RMP) for the quantification of primidone in human serum and plasma
  15. An isotope dilution-liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS)-based candidate reference measurement procedure for the quantification of carbamazepine-10,11-epoxide in human serum and plasma
  16. Should we depend on reference intervals from manufacturer package inserts? Comparing TSH and FT4 reference intervals from four manufacturers with results from modern indirect methods and the direct method
  17. Comparison of three chatbots as an assistant for problem-solving in clinical laboratory
  18. Evidence-based cutoffs for total and adjusted calcium: a major factor in detecting severe hypo- and hypercalcemia
  19. Minor head injury in anticoagulated patients: performance of biomarkers S100B, NSE, GFAP, UCH-L1 and Alinity TBI in the detection of intracranial injury. A prospective observational study
  20. A comparative evaluation of the analytical performances of premier resolution-high-performance liquid chromatography (PR-HPLC) with capillary zone electrophoresis (CZE) assays for the detection of hemoglobin variants and the quantitation of HbA0, A2, E, and F
  21. Get reliable laboratory findings – how to recognize the deceptive effects of angiotensin-converting enzyme inhibitor therapy in the laboratory diagnostics of sarcoidosis?
  22. Reference Values and Biological Variations
  23. Vitamin D and vitamin K status in postmenopausal women with normal and low bone mineral density
  24. Hematology and Coagulation
  25. An automatic analysis and quality assurance method for lymphocyte subset identification
  26. Cancer Diagnostics
  27. Machine learning-based delta check method for detecting misidentification errors in tumor marker tests
  28. Cardiovascular Diseases
  29. Analytical evaluation of the novel Mindray high sensitivity cardiac troponin I immunoassay on CL-1200i
  30. Infectious Diseases
  31. A reactive monocyte subset characterized by low expression of CD91 is expanded during sterile and septic inflammation
  32. Letters to the Editor
  33. Inadvertent omission of a specimen integrity comment – an overlooked post-analytical error
  34. Falsely elevated T3 due to interference of anti-T3 autoantibodies: a case report
  35. Validation of the Siemens Atellica cortisol immunoassay compared to liquid chromatography mass spectrometry in adrenal venous sampling for primary hyperaldosteronism
  36. Lessons learned from site-specific sampling and biological half-life of IGFII and IIE(68-88) peptide: a case study
  37. The added value of automated HPC count: detecting clinically important interferences on the flow cytometric CD34+ cell count
  38. Clinical pilot study on microfluidic automation of IGH-VJ library preparation for next generation sequencing
  39. Long-term effects of interventions applied to optimize the use of 25-OH vitamin D tests in primary health care
https://doi.org/10.1515/cclm-2023-1458
Schlagwörter für diesen Artikel
capillary zone electrophoresis; hemoglobin analysis; hemoglobinopathy; high-performance liquid chromatography; premier resolution; thalassemia

Artikel in diesem Heft

  1. Frontmatter
  2. Editorial
  3. LC-MS/MS random access automation – a game changer for the 24/7 clinical laboratory
  4. Reviews
  5. Neurofilament light protein as a biomarker for spinal muscular atrophy: a review and reference ranges
  6. Differential diagnosis of ascites: etiologies, ascitic fluid analysis, diagnostic algorithm
  7. Opinion Papers
  8. Clinical Decision Support System in laboratory medicine
  9. Blood over-testing: impact, ethical issues and mitigating actions
  10. General Clinical Chemistry and Laboratory Medicine
  11. An isotope dilution-liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS)-based candidate reference measurement procedure for the quantification of zonisamide in human serum and plasma
  12. An isotope dilution-liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS)-based candidate reference measurement procedure for the quantification of carbamazepine in human serum and plasma
  13. An isotope dilution-liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS)-based candidate reference measurement procedure (RMP) for the quantification of phenobarbital in human serum and plasma
  14. An isotope dilution-liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS)-based candidate reference measurement procedure (RMP) for the quantification of primidone in human serum and plasma
  15. An isotope dilution-liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS)-based candidate reference measurement procedure for the quantification of carbamazepine-10,11-epoxide in human serum and plasma
  16. Should we depend on reference intervals from manufacturer package inserts? Comparing TSH and FT4 reference intervals from four manufacturers with results from modern indirect methods and the direct method
  17. Comparison of three chatbots as an assistant for problem-solving in clinical laboratory
  18. Evidence-based cutoffs for total and adjusted calcium: a major factor in detecting severe hypo- and hypercalcemia
  19. Minor head injury in anticoagulated patients: performance of biomarkers S100B, NSE, GFAP, UCH-L1 and Alinity TBI in the detection of intracranial injury. A prospective observational study
  20. A comparative evaluation of the analytical performances of premier resolution-high-performance liquid chromatography (PR-HPLC) with capillary zone electrophoresis (CZE) assays for the detection of hemoglobin variants and the quantitation of HbA0, A2, E, and F
  21. Get reliable laboratory findings – how to recognize the deceptive effects of angiotensin-converting enzyme inhibitor therapy in the laboratory diagnostics of sarcoidosis?
  22. Reference Values and Biological Variations
  23. Vitamin D and vitamin K status in postmenopausal women with normal and low bone mineral density
  24. Hematology and Coagulation
  25. An automatic analysis and quality assurance method for lymphocyte subset identification
  26. Cancer Diagnostics
  27. Machine learning-based delta check method for detecting misidentification errors in tumor marker tests
  28. Cardiovascular Diseases
  29. Analytical evaluation of the novel Mindray high sensitivity cardiac troponin I immunoassay on CL-1200i
  30. Infectious Diseases
  31. A reactive monocyte subset characterized by low expression of CD91 is expanded during sterile and septic inflammation
  32. Letters to the Editor
  33. Inadvertent omission of a specimen integrity comment – an overlooked post-analytical error
  34. Falsely elevated T3 due to interference of anti-T3 autoantibodies: a case report
  35. Validation of the Siemens Atellica cortisol immunoassay compared to liquid chromatography mass spectrometry in adrenal venous sampling for primary hyperaldosteronism
  36. Lessons learned from site-specific sampling and biological half-life of IGFII and IIE(68-88) peptide: a case study
  37. The added value of automated HPC count: detecting clinically important interferences on the flow cytometric CD34+ cell count
  38. Clinical pilot study on microfluidic automation of IGH-VJ library preparation for next generation sequencing
  39. Long-term effects of interventions applied to optimize the use of 25-OH vitamin D tests in primary health care
Jetzt anmelden und 20% sparen
Institutioneller Zugang
Wie funktioniert Authentifizierung?
Haben Sie eine Idee, wie wir unsere Website verbessern können?
Bitte schreiben Sie uns.
© 2026 De Gruyter Brill
Heruntergeladen am 20.1.2026 von https://www.degruyterbrill.com/document/doi/10.1515/cclm-2023-1458/html
Button zum nach oben scrollen