Development of hemoglobin typing control materials for laboratory investigation of thalassemia and hemoglobinopathies

Sakorn Pornprasert; Monthathip Tookjai; Manoo Punyamung; Panida Pongpunyayuen; Kanokwan Jaiping

doi:10.1515/cclm-2015-0113

Article

Development of hemoglobin typing control materials for laboratory investigation of thalassemia and hemoglobinopathies

Sakorn Pornprasert , Monthathip Tookjai , Manoo Punyamung , Panida Pongpunyayuen and Kanokwan Jaiping

Published/Copyright: May 21, 2015

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From the journal Clinical Chemistry and Laboratory Medicine (CCLM) Volume 54 Issue 1

Abstract

Background: To date, the hemoglobin (Hb) typing control materials for laboratory investigation of thalassemia with low (1.8%–3.2%) and high (4%–6%) levels of HbA₂ are available but there are no Hb typing quality control materials for analysis of thalassemia and hemoglobinopathies which are highly prevalent in South-East Asian countries. The main aim of the present study was to develop the lyophilized Hb typing control materials for laboratory investigation of thalassemia and hemoglobinopathies that are commonly found in South-East Asia.

Methods: Erythrocytes of blood samples containing Hb Bart’s, HbH, HbE, HbF, Hb Constant Spring (CS), Hb Hope, and Hb Q-Thailand were washed and dialysed with 0.85% saline solution. The erythrocytes were then lysed in 5% sucrose solution. The lyophilized Hb typing control materials were prepared by using a freeze drying (lyophilization) method. The high performance liquid chromatography (HPLC) analysis of lyophilized Hb was performed after the storage at −20 °C for 1 year and also after reconstitution and storage at 4 or −20 °C for 30 days. In addition, the Hb analysis was compared between the three different methods of HPLC, low pressure liquid chromatography (LPLC) and capillary electrophoresis (CE).

Results: Following a year of storage at −20 °C, the HPLC chromatograms of lyophilized Hb typing control materials showed similar patterns to the equivalent fresh whole blood. The stability of reconstituted Hb typing control materials was also observed through 30 days after reconstitution and storage at −20 °C. Moreover, the Hb typing control materials could be analyzed by three methods, HPLC, LPLC and CE. Even a degraded peak of HbCS was found on CE electropherogram.

Conclusions: The lyophilized Hb typing control materials could be developed and used as control materials for investigation of thalassemia and hemoglobinopathies.

Keywords: capillary electrophoresis; control materials; hemoglobin analysis; hemoglobinopathy; HPLC

Corresponding author: Dr. Sakorn Pornprasert, Faculty of Associated Medical Sciences, Department of Medical Technology, Chiang Mai University, 110 Intawaroros Road, Chiang Mai, 50200 Thailand, Phone: +66 53 945078, Fax: +66 53 946042, E-mail: sakornmi001@gmail.com

Acknowledgments

The authors thank technicians of the Associated Medical Sciences Clinical Service Center, Faculty of Associated Medical Sciences, Chiang Mai University, Chiang Mai, Thailand and the technicians in the Medical Technology Department of Lamphun Hospital, Lamphun, Thailand for their help and assistance. We also would like to express our gratitude to Roscoe C Butler Jr and Prof. Dr. Srijit Das for editing the manuscript. This study was supported by grants from the National Research Council of Thailand (NRCT), Thailand.

Author contributions: All the authors have accepted responsibility for the entire content of this submitted manuscript and approved submission.
Research funding: None declared.
Employment or leadership: None declared.
Honorarium: None declared.
Competing interests: The funding organization(s) played no role in the study design; in the collection, analysis, and interpretation of data; in the writing of the report; or in the decision to submit the report for publication.

References

1. Fucharoen S, Winichagoon P. Thalassemia in SouthEast Asia: problems and strategy for prevention and control. Southeast Asian J Trop Med Public Health 1992;23:647–55.Search in Google Scholar

2. Fucharoen S, Winichagoon P. Hemoglobinopathies in Southeast Asia: molecular biology and clinical medicine. Hemoglobin 1997;21:299–319.10.3109/03630269709000664Search in Google Scholar PubMed

3. Tanphaichitr VS, Mahasandana C, Suvatte V, Yodthong S, Pung-amritt P, Seeloem J. Prevalence of hemoglobin E, alpha-thalassemia and glucose-6-phosphate dehydrogenase deficiency in 1,000 cord bloods studied in Bangkok. Southeast Asian J Trop Med Public Health 1995;26 (Suppl 1):271–4.Search in Google Scholar

4. Tienthavorn V, Pattanapongsthorn J, Charoensak S, Sae-Tung R, Charoenkwan P, Sanguansermsri T. Prevalence of thalassemia carriers in Thailand. Thai J Hematol Transfus Med 2006;16:307–12.Search in Google Scholar

5. Charoenkwan P, Taweephol R, Sirichotiyakul S, Tantiprabha W, Sae-Tung R, Suanta S, et al. Cord blood screening for alpha-thalassemia and hemoglobin variants by isoelectric focusing in northern Thai neonates: correlation with genotypes and hematologic parameters. Blood Cells Mol Dis 2010;45:53–7.10.1016/j.bcmd.2010.02.015Search in Google Scholar PubMed

6. Sura T, Busabaratana M, Youngcharoen S, Wisedpanichkij R, Viprakasit V, Trachoo O. Haemoglobin Hope in a northern Thai family: first identification of homozygous haemoglobin hope associated with haemoglobin H disease. Eur J Haematol 2007;79:251–4.10.1111/j.1600-0609.2007.00907.xSearch in Google Scholar PubMed

7. Sirichotiyakul S, Tantipalakorn C, Sanguansermsri T, Wanapirak C, Tongsong T. Erythrocyte osmotic fragility test for screening of alpha-thalassemia-1 and beta- thalassemia trait in pregnancy. Int J Gynaecol Obstet 2004;86: 347–50.10.1016/j.ijgo.2004.04.037Search in Google Scholar PubMed

8. Nathalang O, Nillakupt K, Arnutti P, Boonsiri T, Panichkul S, Areekul W. Screening for thalassemia and hemoglobinopathy in a rural area of Thailand: a preliminary study. J Med Assoc Thai 2005;88(Suppl 3):S35–42.Search in Google Scholar

9. Tatsumi N, Tsuda I, Funahara Y, Bunyaratvej A, Fucharoen S. Automatic measurement of hemoglobin F in blood obtained from patients with hemoglobin E/E and beta-thalassemia/hemoglobin/E. Southeast Asian J Trop Med Public Health 1992;23(Suppl 2):91–4.Search in Google Scholar

10. Tatsumi N, Tsuda I, Funahara Y, Bunyaratvej A, Pootrakul P, Fucharoen S. Analysis of hematological data of thalassemia cases in Thailand. Nippon Ketsueki Gakkai Zasshi 1989;52:792–5.Search in Google Scholar

11. 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: 484–95.10.1111/j.1751-553X.2009.01158.xSearch in Google Scholar PubMed

12. Panyasai S, Sringam P, Fucharoen G, Sanchaisuriya K, Fucharoen S. A simplified screening for alpha-thalassemia 1 (SEA type) using a combination of a modified osmotic fragility test and a direct PCR on whole blood cell lysates. Acta Haematol 2002;108:74–8.10.1159/000064746Search in Google Scholar PubMed

13. Sanguansermsri T, Thanarattanakorn P, Steger HF, Tongsong T, Chanprapaph P, Wanpirak C, et al. Prenatal diagnosis of beta-thalassemia major by high-performance liquid chromatography analysis of hemoglobins in fetal blood samples. Hemoglobin 2001;25:19–27.10.1081/HEM-100103066Search in Google Scholar PubMed

14. Pornprasert S, Kongthai K, Waneesorn J, Jaiping K, Treesuwan K. Development of control material for hemoglobin analysis. Clin Chem Lab Med 2012;50:351–6.10.1515/cclm.2011.777Search in Google Scholar PubMed

15. Pornprasert S, Phusua A, Suanta S, Saetung R, Sanguansermsri T. Detection of alpha-thalassemia-1 Southeast Asian type using real-time gap-PCR with SYBR Green1 and high resolution melting analysis. Eur J Haematol 2008;80:510–4.10.1111/j.1600-0609.2008.01055.xSearch in Google Scholar PubMed

16. Pornprasert S, Punyamung M, Treesuwan K. Criteria for detection of alpha-thalassemia-1 Thai type deletion in routine laboratory. Clin Lab 2013;59:1423–7.10.7754/Clin.Lab.2013.121217Search in Google Scholar

17. Paleari R, Munoz A, Mosca A. Towards the development of a certified reference material for hemoglobin A2. Clin Chem Lab Med 2010;48:1611–8.10.1515/CCLM.2010.317Search in Google Scholar PubMed

18. Hunt DM, Higgs DR, Winichagoon P, Clegg JB, Weatherall DJ. Haemoglobin Constant Spring has an unstable alpha chain messenger RNA. Br J Haematol 1982;51:405–13.10.1111/j.1365-2141.1982.tb02796.xSearch in Google Scholar PubMed

19. Pornprasert S, Panyasai S, Waneesorn J, Kongthai K, Singboottra P. Quantification of hemoglobin Constant Spring in heterozygote and homozygote by a capillary electrophoresis method. Int J Lab Hematol 2012;34:143–7.10.1111/j.1751-553X.2011.01371.xSearch in Google Scholar PubMed

20. Rodak BF, Coleman M. Hemoglobin Metabolism. In: Rodak BF, editor. Hematology clinical principles and applications. Philadephia: WB Saunders, 2002:105–14.Search in Google Scholar

Received: 2015-2-2

Accepted: 2015-4-20

Published Online: 2015-5-21

Published in Print: 2016-1-1

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Articles in the same Issue

https://doi.org/10.1515/cclm-2015-0113

Keywords for this article

capillary electrophoresis; control materials; hemoglobin analysis; hemoglobinopathy; HPLC