Monoclonal antibodies against hemoglobins for detecting thalassemia
Article Sidebar
Main Article Content
Abstract
Background: Hemoglobin is composed of globin polypeptide chains, which serve as immunogens to induce the production of antibodies.
Objectives: This review article aims to describe the use of antibodies against human hemoglobins for the identification of thalassemia and hemoglobinopathies.
Materials and methods: Literature review to discusses the nature of normal human hemoglobin, hemoglobin switching, thalassemia and hemoglobinopathies, laboratory diagnosis, general properties of antibodies, production of antibodies against human hemoglobins, and clinical applications of these antibodies in identifying thalassemia and hemoglobinopathies.Antibody-based detection of hemoglobin is highly useful in diagnosing thalassemia and hemoglobinopathies.
Results: Polyclonal antibodies against HbF have been applied in sandwich ELISA to accurately detect HbF levels. Monoclonal antibodies against HbH and Hb Bart’s have been produced and utilized in sandwich ELISA for detecting α-thalassemia. In addition, monoclonal antibodies against hemoglobin containing α-globin chains were developed and applied in sandwich ELISA to identify infants with Hb Bart’s hydrops fetalis, a condition in which no α-globin chains are produced. For detecting β-thalassemia carriers, monoclonal antibodies against HbA2 were produced, and sandwich ELISA was employed to measure HbA2 levels, which are elevated in these individuals.
Conclusion: Antibody-based diagnosis of thalassemia and hemoglobinopathies enhances the quality of screening platforms and makes diagnosis of these disorders more reliable.
Article Details
This work is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.
Personal views expressed by the contributors in their articles are not necessarily those of the Journal of Associated Medical Sciences, Faculty of Associated Medical Sciences, Chiang Mai University.
References
Perutz M. Hemoglobin structure. In: Bunn FF, Forget BG, eds. Hemoglobin: Molecular genetics and clinical aspects. Philadelphia: WB Sauners; 1986:13-35.
Bunn HF, Forget BG, Ranney HM. Human Hemoglobins. Toronto: W.B. Saunders Company; 1977.
Weatherall DJ, Clegg JB. The Thalassaemia Syndromes (4th Ed). Oxford: Blackwell Scientific; 2001.
Wilber A, Nienhuis AW, Persons DA. Transcriptional regulation of fetal to adult hemoglobin switching: New therapeutic opportunities. Blood. 2011; 117(15): 3945-53. doi: 10.1182/blood-2010-11-316893.
Fucharoen S, Winichagoon P. Thalassemia in SouthEast Asia: problems and strategy for prevention and control. Southeast Asian J Trop Med Public Health. 1992; 23(4): 647-55. PMID: 1298071.
Fucharoen S, Winichagoon P. Hemoglobinopathies in Southeast Asia: molecular biology and clinical medicine. Hemoglobin. 1997; 21(4): 299-319. doi: 10.3109/03630269709000664.
Harteveld CL, Higgs DR. Alpha-thalassaemia. Orphanet J Rare Dis. 2010; 5: 13. Doi: 10.1186/1750-1172-5-13
Thein SL. The molecular basis of beta-thalassemia. Cold Spring Harb Perspect Med. 2013; 3(5): a011700. doi: 10.1101/cshperspect.a011700.
Thom CS, Dickson CF, Gell DA, Weiss MJ. Hemoglobin variants: biochemical properties and clinical correlates. Cold Spring Harb Perspect Med. 2013; 3(3): a011858. doi: 10.1101/cshperspect. a011858.
Laboratories of Computer Science & Engineering and Biochemistry & Molecular Biology at the Pennsylvania State University. Globin Gene Server. 2022; https://globin.bx.psu.edu/.
Charoenkwan P, Taweephon R, Sae-Tung R, Thanarattanakorn P, Sanguansermsri T. Molecular and clinical features of Hb H disease in northern Thailand. Hemoglobin. 2005; 29(2): 133-140. PMID: 15921165.
Fucharoen S, Viprakasit V. Hb H disease: clinical course and disease modifiers. Hematology Am Soc Hematol Educ Program. 2009: 26-34. doi: 10.1182/asheducation-2009.1.26.
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(3): 405-13. PMID: 7104225.
Laig M, Pape M, Hundrieser J, et al. The distribution of the Hb constant spring gene in Southeast Asian populations. Hum Genet. 1990; 84(2): 188-90. doi: 10.1007/BF00208939.
Laosombat V, Viprakasit V, Chotsampancharoen T, et al. Clinical features and molecular analysis in Thai patients with HbH disease. Ann Hematol. 2009; 88(12): 1185-92. doi: 10.1007/s00277-009-0743-5.
Old JM. Screening and genetic diagnosis of haemoglobin disorders. Blood Rev. 2003; 17(1): 43-53. doi: 10.1016/s0268-960x(02)00061-9.
Fucharoen S, Winichagoon P, Piankijagum A. Standardization on laboratory diagnosis of thalassemia and abnormal hemoglobin. Southeast Asian J Trop Med Public Health. 1999; 30) Suppl 3): 90-8. PMID: 10926267.
Leckngam P, Limweeraprajak E, Kiewkarnkha T, Tatu T. The Hb E (HBB: c.79G>A), Mean corpuscular volume, mean corpuscular hemoglobin cutoff points in double heterozygous Hb E/- -(SEA) alphathalassemia-1 carriers are dependent on hemoglobin levels. Hemoglobin. 2017; 41(1): 38-43. doi: 10.1080/03630269.2017.1295984.
Manglani M, Lokeshwar MR, Vani VG, Bhatia N, Mhaskar V. ‘NESTROFT’--an effective screening test for beta thalassemia trait. Indian pediatrics. 1997; 34(8): 702-7. PMID: 9492398.
Munkongdee T, Chen P, Winichagoon P, Fucharoen S, Paiboonsukwong K. Update in Laboratory Diagnosis of Thalassemia. Front Mol Biosci. 2020; 7: 74. doi: 10.3389/fmolb.2020.00074.
Sanchaisuriya K, Fucharoen S, Fucharoen G, et al. A reliable screening protocol for thalassemia and hemoglobinopathies in pregnancy: an alternative approach to electronic blood cell counting. Am J Clin Pathol. 2005; 123(1): 113-8. doi: 10.1309/fuf9evgq24v1pktp.
Sanguansermsri T, Phumyu N, Chomchuen S, Steger HF. Screening for alpha-thalassemia-1 heterozygotes in expecting couples by the combination of a simple erythrocyte osmotic fragility test and a PCR-based method. Community Genet. 1999; 2(1): 26-9. doi: 10.1159/000016180.
Sirichotiyakul S, Maneerat J, Sa-nguansermsri T, Dhananjayanonda P, Tongsong T. Sensitivity and specificity of mean corpuscular volume testing for screening for alpha-thalassemia-1 and betathalassemia traits. J Obstet Gynaecol Res. 2005; 31(3): 198-201. doi: 10.1111/j.1447-0756.2005.00280.x.
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(3): 347-350. doi: 10.1016/j.ijgo.2004.04.037.
Sirichotiyakul S, Tongprasert F, Tongsong T. Screening for hemoglobin E trait in pregnant women. Int J Gynaecol Obstet. 2004; 86(3): 390-1. doi: 10.1016/j.ijgo.2004.04.041.
Janeway CA, Travers P, Walport M. The structure of typical antibody molecule. In: Immunology: The Immune System in Heath and Disease 5th ed. New York: Garland Science; 2001.
Feng L, Wang X, Jin H. Rabbit monoclonal antibody: potential application in cancer therapy. Am J Transl Res. 2011; 3(3): 269-274. PMID: 21633632.
Rossi S, Laurino L, Furlanetto A, et al. Rabbit monoclonal antibodies: a comparative study between a novel category of immunoreagents and the corresponding mouse monoclonal antibodies. Am J Clin Pathol. 2005; 124(2): 295-302. doi: 10.1309/NR8H-N08G-DPVE-MU08.
Actor JK. Assessment of immune parameters and immunodiagnostics. In: Actor JK, ed. Introductory Immunology; Basic Concepts for Interdisciplinary Applications (3rd Ed). Academic Press; 2023:169-188. doi: 10.1016/B978-0-12-420030-2.00019-6
Kerdpoo S, Laopajon W, Kasinrerk W, Pata S, Tatu T. A modified sandwich ELISA for accurate measurement of HbF in alpha-thalassemia carriers containing Hb Bart’s and Hb Portland 1. J Immunoassay Immunochem. 2018; 39(3): 323-36. doi: 10.1080/15321819.2018.1488726.
Pakdeepak K, Pata S, Chiampanichayakul S, Kasinrerk W, Tatu T. Production and characterization of monoclonal antibodies against alpha-globin chain-containing human hemoglobins for detecting alpha-thalassemia disease. J Immunoassay Immunochem. 2016; 37(6): 564-71. doi: 10.1080/15321819.2016.1174135.
Tatu T, Kiewkarnkha T, Khuntarak S, Khamrin S, Suwannasin S, Kasinrerk W. Screening for coexistence of alpha-thalassemia in beta-thalassemia and in HbE heterozygotes via an enzyme-linked immunosorbent assay for Hb Bart’s and embryonic zeta-globin chain. Int J Hematol. 2012; 95(4): 386-93. doi: 10.1007/s12185-012-1039-4.
Shyamala M, Kiefer CR, Moscoso H, Garver FA. A monoclonal antibody-linked immunoassay for hemoglobin H disease. Ann Hematol. 1992; 65(1): 37-40. doi: 10.1007/BF01715124.
Garver FA, Singh H, Moscoso H, Kestler DP, McGuire BS, Jr. Identification and quantification of hemoglobins A2 and Barts with an enzyme-labeled immunosorbent assay. Clin Chem. 1984; 30(7): 1205-8. PMID: 6203666.
Tayapiwatana C, Kuntaruk S, Tatu T, et al. Simple method for screening of alpha-thalassaemia 1 carriers. Int J Hematol. 2009; 89(5): 559-67. doi: 10.1007/s12185-009-0331-4.
Panyasai S, Permsripong N, Jaiping K, Khantarag P, Pornprasert S. Hematological and Molecular Characterization of Hb J-Buda [alpha61(E10) Lys --> Asn, AAG > AAT]. Indian J Hematol Blood Transfus. 2016; 32(Suppl 1): 311-4. doi: 10.1007/s12288-016-0641-7.
Jiang F, Ju AP, Li J, et al. Hb Westmead (HBA2:c.369C>G): Hematological characteristics in heterozygotes with and without alpha(0)-thalassemia. Hemoglobin. 2020; 44(3): 153-5. doi: 10.1080/03630269.2020.1768109.
Panyasai S, Jaiping K, Khantarag P, Nochod P, Satthakarn S. Unraveling thalassemia intermedia: Novel insights of a hemoglobin Jax [HBA2:c.44G>C] and deletional alpha(0)-thalassemia interaction phenotype. Hematol Transfus Cell Ther. 2025; 47(1): 103739. doi: 10.1016/j.htct.2025.103739.
Chan G, Wong J, Tan T, Ghallyan N, Browett P. False positive Hb Bart’s immunochromatographic strip test for alpha thalassaemia in delta-beta thalassaemia carriers. Int J Lab Hematol. 2023; 45(6): 984-6. doi: 10.1111/ijlh.14122.
Theingi M, Khamphikham P, Pornprasert S. Consideration of immunochromatographic strip positive results for screening of alpha-thalassemia in elevated hemoglobin F samples. Scand J Clin Lab Invest. 2025; 85(5): 346-52. doi: 10.1080/00365513.2025.2524849.
Wanapirak C, Piyamongkol W, Sirichotiyakul S, Tayapiwatana C, Kasinrerk W, Tongsong T. Accuracy of immunochromatographic strip test in diagnosis of alpha-thalassemia-1 carrier. J Med Assoc Thai. 2011; 94(7): 761-5. PMID: 21774279.
Prayalaw P, Fucharoen G, Fucharoen S. Routine screening for alpha-thalassaemia using an immunochromatographic strip assay for haemoglobin Bart’s. J Med Screen. 2014; 21(3): 120-5. doi: 10.1177/0969141314538611.
Sudjaroen Y. Efficiency assessment of immunochromatographic strip test for the diagnosis of alpha-thalassemia-1 carriers. J Lab Physicians. 2015; 7(1): 4-10. doi: 10.4103/0974-2727.154779.
Winichagoon P, Kumpan P, Holmes P, et al. Validation of the immunochromatographic strip for alpha-thalassemia screening: a multicenter study. Transl Res. 2015; 165(6): 689-95. doi: 10.1016/j.trsl.2014.10.013.
Bunkall C, Ghallyan N, Elliott C, Van de Water N,Chan G. Evaluation of an immunochromatographic strip test for alpha-thalassaemia screening. Int J Lab Hematol. 2018; 40(6): 691-6. doi: 10.1111/ijlh.12905.
Luo HY, Clarke BJ, Gauldie J, Patterson M, Liao SK, Chui DH. A novel monoclonal antibody based diagnostic test for alpha-thalassemia-1 carriers due to the (-SEA/) deletion. Blood. 1988; 72(5): 1589-94. PMID: 3179441.
Chui DH, Luo HY, Clarke BJ. Potential application of a new screening test for alpha-thalassemia-1 carriers. Hemoglobin. 1988; 12(5-6): 459-63. doi: 10.3109/03630268808991635.[48] Ireland JH, Luo HY, Chui DH, Chu B, Yuen J, Hsia YE. Detection of the (--SEA) double alpha-globin gene deletion by a simple immunologic assay for embryonic zeta-globin chains. Am J Hematol. 1993; 44(1): 22-8. doi: 10.1002/ajh.2830440106.
Lafferty JD, Crowther MA, Waye JS, Chui DH. A reliable screening test to identify adult carriers of the (--SEA) alpha zero-thalassemia deletion. Detection of embryonic zeta-globin chains by enzyme-linked immunosorbent assay. Am J Clin Pathol. 2000;114(6):927-931. doi: 10.1309/26G7-BQH4-93BV-UR0Q.
Pata S, Khummuang S, Pornprasert S, Tatu T, Kasinrerk W. A simple and highly sensitive ELISA for screening of the alpha-thalassemia-1 Southeast Asian-type deletion. J Immunoassay Immunochem. 2014; 35(2): 194-206. doi: 10.1002/ajh.2830380311.
Pata S, Pongpaiboon M, Laopajon W, et al. Immunostick test for detecting zeta-globin chains and screening of the Southeast Asian alphathalassemia 1 Deletion. Biol Proceed Online. 2019; 21: 15. doi: 10.1186/s12575-019-0104-2.
Pata S, Laopajon W, Pongpaiboon M, et al. Impact of the detection of zeta-globin chains and hemoglobin Bart’s using immunochromatographic strip tests for alpha0-thalassemia (--SEA) differential diagnosis. PLoS One. 2019; 14(10): e0223996. doi: 10.1371/journal.pone.0223996
Shyamala M, Kiefer CR, Moscoso H, Garver FA. Application of a monoclonal antibody specific for the delta chain of hemoglobin A2 in the diagnosis of beta thalassemia. Am J Hematol. 1991; 38(3): 214-9. doi: 10.1002/ajh.2830380311.
Kuntaruk S, Tatu T, Keowkarnkah T, Kasinrerk W. Sandwich ELISA for hemoglobin A2 quantification and identification of beta-thalassemia carriers. Int J Hematol. 2010; 91(2): 219-28. doi: 10.1007/s12185-009-0490-3.
Committee for Developing Direction of Laboratory Diagnosis of Thalassemia and Hemoglobinopathies, Clinical Research Center, Department of Medical Sciences, Ministry of Public Health. Direction of Laboratory Diagnosis of Thalassemia and Hemoglobinopathies. Nonthaburi, Mudded Ltd. Co., 2009 (in Thai).
Colaco S, Colah R, Nadkarni A. Significance of borderline HbA(2) levels in beta thalassemia carrier screening. Sci Rep. 2022; 12(1): 5414. doi: 10.1038/s41598-022-09250-5
