Alpha Thalassemia Trait Diagnosis: Comprehensive Guide for Healthcare Professionals

Introduction

Hemoglobin, the vital protein in red blood cells, is responsible for oxygen transport throughout the body. It comprises alpha (α) and beta (β) globin chains, crucial for erythrocyte function and shape. Disruptions in globin chain production lead to hemoglobinopathies, with thalassemia representing quantitative defects. Alpha (α)-thalassemia arises from reduced or absent production of α-globin chains, while beta (β)-thalassemia involves β-globin chains.

Alpha-thalassemia is a prevalent genetic blood disorder globally. The production of α-globin is governed by four α genes located on chromosome 16. Alpha-thalassemia occurs when there is a reduction (α+) or complete absence (α°) of globin chain production from these genes. Understanding Alpha Thal Trait Diagnosis is critical because it represents the carrier state of this condition, often presenting without symptoms but significant for genetic counseling.

The carrier state in α-thalassemia can be either α+ trait (α-thalassemia 2, resulting from one α-globin gene deletion) or α° trait (α-thalassemia 1, resulting from two α-globin gene deletions). While α-thalassemia 2 is typically asymptomatic, recognizing both traits is vital for preventing more severe forms like Hemoglobin H disease (HbH, deletion of three α-globin genes) and α-thalassemia major or Hemoglobin Bart’s (Hb Bart’s, deletion of all four α-globin genes), the latter being frequently fatal in utero causing hydrops fetalis.

Differentiating thalassemia from iron deficiency anemia (IDA) is crucial in patients presenting with low hemoglobin levels. Laboratory evaluations, including complete blood count (CBC) and hemoglobin analysis via high-performance liquid chromatography (HPLC) or electrophoresis, are essential for identifying hemoglobin disorders in individuals with chronic anemia. Advanced molecular techniques like allele-specific polymerase chain reaction (PCR), reverse dot blot (RDB) analysis, real-time PCR, and DNA sequencing are valuable for precise genetic diagnosis, especially for alpha thal trait diagnosis and subsequent genetic counseling. Effective management, parental counseling, antenatal diagnosis, newborn screening, and complication prevention are paramount for improving the quality of life for affected individuals and their families.

Types of Alpha-Thalassemia and the Alpha Thal Trait

Alpha-thalassemia is categorized into four main types, based on the number of functional α-globin genes inherited, with symptom severity generally increasing with the number of affected genes. Understanding these types is crucial for accurate alpha thal trait diagnosis and risk assessment.

• Silent Carrier State: Individuals with three functional α-globin genes are silent carriers. They usually show no clinical symptoms and have normal hematological parameters. Diagnosis is typically made through genetic testing of family members of affected individuals.

• Alpha-Thalassemia Trait (Minor) – α-Thalassemia 2 (αα/–) or α-Thalassemia 1 (α-/α-): This condition occurs when two α genes are deleted. α-globin chain production is reduced by approximately half. Adults often compensate with increased red blood cell (RBC) production, maintaining a balance between α- and β-globin chains. Patients are usually asymptomatic or present with mild microcytic anemia. This is the primary focus of alpha thal trait diagnosis.

• Hemoglobin H Disease (HbH) (– -/αα or – -/–α): Resulting from three α genes being deleted, HbH disease leads to a more significant reduction in α-globin production. The excess β-globin chains form tetramers (HbH), which are unstable and precipitate within red cells, leading to hemolytic anemia. Clinical presentation varies from mild to moderate anemia, often requiring medical intervention.

• Alpha-Thalassemia Major (Hydrops Fetalis) (–/–): This is the most severe and often fatal form, occurring when all four α genes are deleted. The complete absence of α-globin chains leads to a predominance of γ-globin tetramers (Hb Bart’s) in fetal life. Hb Bart’s has a very high oxygen affinity, poorly releasing oxygen to tissues, resulting in severe fetal anemia, hydrops fetalis, and typically intrauterine death or neonatal death shortly after birth.

Etiology and Epidemiology of Alpha-Thalassemia

Alpha-globin genes are located on chromosome 16, with individuals typically inheriting four α genes (αα/αα), two from each parent. Alpha-thalassemia arises from deletions in these α-globin genes. The severity of anemia is directly related to the number of deleted genes, influencing the amount of α-globin chain production.

Thalassemia is genetically diverse, with over 200 mutations associated with α-thalassemia. It is most prevalent in Southeast and South Asia, the Middle East, Mediterranean regions, and North and Central Africa. Global migration has increased thalassemia incidence in Northern Europe and North America. Hb Bart’s hydrops fetalis is particularly common in Southeast Asia and Southern China. Understanding the epidemiology is important for targeted alpha thal trait diagnosis and screening programs in at-risk populations.

Pathophysiology of Alpha-Thalassemia and Implications for Alpha Thal Trait

The primary pathophysiological feature in thalassemia is imbalanced globin chain production, leading to fragile RBCs that are easily destroyed in the bone marrow or peripheral blood. This results in chronic anemia, splenomegaly, and skeletal deformities in more severe forms.

In alpha thal trait, the blood picture often mimics iron deficiency anemia (IDA) with slightly microcytic RBCs. However, in the silent carrier type, erythrocytes may be normal. Alpha-thalassemia 1 trait typically presents with mild anemia, slightly decreased RBC indices (MCV and MCH), hypochromia, microcytosis, and anisopoikilocytosis. HbA2 levels are in the low to low-normal range (1.5%-2.5%). Neonates with alpha thal trait may show moderate amounts of Hb Bart’s (3% to 8%) on blood films.

HbH disease shows significantly reduced α-globin synthesis. HbH, composed of β-globin chain homotetramers, is detectable by HPLC or electrophoresis, typically ranging from 3% to 30%. This is associated with mild to severe microcytic or normocytic anemia and elevated bilirubin levels due to hemolysis.

Alpha-thalassemia major (Hb Bart’s hydrops fetalis syndrome) is the most severe form, where no α-globin chains are synthesized. Fetal blood shows only Hb Bart’s (γ4) and some embryonic Hemoglobin Portland. Prenatal diagnosis is crucial for parental counseling in these cases.

Transfusion-dependent thalassemia (TDT) patients develop complications from systemic iron overload due to hemoglobin denaturation, ineffective erythropoiesis, and increased gastrointestinal iron absorption, leading to hemosiderosis and organ damage. Iron chelation therapy is essential for these patients.

Specimen Requirements and Procedures for Alpha Thal Trait Diagnosis

Specimen collection for alpha thal trait diagnosis, as well as other forms of alpha-thalassemia, depends on the diagnostic tests chosen. A range of laboratory tests is required, starting with a CBC. Hemoglobin analysis identifies and quantifies hemoglobin types, including HbH and Hb Bart’s. EDTA vials are used for whole blood collection for CBC, hemoglobin analysis, and molecular testing.

For prenatal alpha thal trait diagnosis in suspected carriers or parents with thalassemia minor, cytogenetic analysis on chorionic villi samples or amniotic fluid cells is preferred. Chorionic villi can be preserved in culture medium for up to 7 days.

Diagnostic Tests for Alpha Thal Trait and Alpha-Thalassemia

Initial diagnostic tests for alpha thal trait diagnosis and other α-thalassemia types include:

• Complete Blood Count (CBC): Automated hematology analyzers are used for CBC. While hemoglobin level, MCV, and MCH alone are not sufficient to differentiate thalassemia trait from IDA or α- from β-thalassemia, an elevated RBC count can help distinguish α-thalassemia from IDA, which often presents with a low RBC count. Platelet and WBC counts are typically unaffected in thalassemia.

• Iron Studies: Serum ferritin and transferrin levels are crucial to differentiate thalassemia from IDA. Normal or slightly increased ferritin with near-normal transferrin is more consistent with thalassemia, whereas IDA typically shows low ferritin and elevated transferrin.

• Peripheral Blood Smear: In thalassemia, peripheral smears may show microcytic, hypochromic anemia, target cells, teardrop cells, and basophilic stippling, although these are also seen in IDA. HbH disease is indicated by golf-ball-like hemoglobin inclusions.

• Hemoglobin Analysis (HPLC or Electrophoresis): These techniques quantify different hemoglobin types in blood, crucial for differentiating thalassemia types, including alpha thal trait. They offer precision and reproducibility.

• Molecular Testing (allele-specific PCR, RDB analysis, gap-PCR, real-time PCR, DNA sequencing): Molecular tests identify the specific mutations causing α-thalassemia, essential for accurate alpha thal trait diagnosis, genetic counseling, and research.

Interpretation of test results must be combined with clinical findings to guide management decisions.

Testing Procedures for Alpha-Thalassemia Diagnosis

High-throughput techniques are advantageous for speed, efficiency, and scalability in detecting and characterizing α-thalassemia mutations. The choice of technique depends on resolution needs and available resources.

• High-Performance Liquid Chromatography (HPLC): HPLC separates, identifies, and quantifies hemoglobin molecules using a slightly negatively charged silica gel column. Hemoglobin variants are eluted based on their affinity for the column using inorganic phosphate buffers. Chromatograms show retention time, peaks, and percentages of hemoglobin fractions for accurate diagnosis of hemoglobinopathies and thalassemia, including identifying patterns suggestive of alpha thal trait.

• Capillary Electrophoresis System: Hemoglobin components are separated in silica capillaries using an alkaline buffer medium. Hemoglobin migrates to the anode due to its negative charge in alkaline conditions. Structural variants separate based on surface charge differences, and photometry at 415 nm quantifies each variant. Capillary electrophoresis is useful for both prenatal and postnatal hemoglobinopathy diagnosis and can detect HbH, Hb Bart’s, and HbCS, important markers in more severe forms but also relevant in excluding them in alpha thal trait diagnosis.

• Allele-Specific Polymerase Chain Reaction (PCR): PCR uses primers with sequences identical except at the 3′-terminus base. One primer matches the wild-type, and the other the mutant base. Taq polymerase amplifies genes only with perfect primer-template matching. In alpha thal trait diagnosis, this can identify specific gene deletions or mutations.

• Reverse Dot Blot Analysis: PCR products are dotted on a membrane filter and hybridized with allele-specific oligomeric DNA probes, often radiolabeled or enzyme-linked. This method facilitates easy identification of suspected mutations related to alpha thal trait and other forms.

• Real-Time PCR with Melting Curve Analysis: Real-time PCR is faster and less labor-intensive than conventional PCR, using fluorescent signals from product synthesis. Dyes like SYBR® Green are used. Melting curve analysis helps differentiate α-thalassemia 1 and α-thalassemia 2 heterozygotes, HbH disease, and α-thalassemia 1 homozygotes (Hb Bart’s), providing a comprehensive tool for alpha thal trait diagnosis and beyond.

• Direct DNA Sequencing (Sanger Sequencing): PCR product sequencing identifies specific gene mutations in α-thalassemia, using Sanger’s dideoxy termination method, the most widely used technique for definitive mutation identification in alpha thal trait cases.

• Multiplex Ligation-Dependent Probe Amplification (MLPA): MLPA uses multiplex PCR to detect deletions or duplications in screened regions, useful for finding known and unknown deletions in unsolved cases when conventional techniques fail, especially important for complex alpha thal trait diagnosis.

• Next-Generation Sequencing (NGS): NGS technology sequences the entire human genome at ultra-high throughput. Targeted NGS can analyze globin gene regions, regulatory regions, and modifier genes. NGS is more accurate than conventional methods, increases sequencing capacity, requires less sample input, and detects variants at lower allele frequencies than Sanger sequencing, offering advanced capabilities for alpha thal trait diagnosis and comprehensive genetic analysis.

Interfering Factors in Alpha-Thalassemia Diagnostic Tests

Accuracy in diagnostic tests for alpha thal trait and other alpha-thalassemia conditions can be compromised by preanalytical, analytical, and postanalytical factors. Preanalytical errors include improper specimen collection, transport, or storage. Hemoglobin denaturation can occur if samples are stored improperly. Poor sample preparation, such as maternal tissue contamination in chorionic villi samples, can also cause errors.

Analytical errors can arise from using improperly stored reagents or buffers. Postanalytical factors, such as rounding off reported figures, can also lead to diagnostic inaccuracies. Accessibility to advanced tests like NGS and allele-specific PCR is limited by high costs and technical demands, impacting widespread use in diverse populations for alpha thal trait diagnosis.

Results, Reporting, and Critical Findings in Alpha Thal Trait Diagnosis

Healthy adults typically have Hemoglobin A (HbA) as the major hemoglobin (95%-98%), with Hemoglobin A2 (HbA2) at 2%-3% and Hemoglobin F (HbF) less than 2%. Individuals with α-thalassemia show varying degrees of anemia, indicated by low hemoglobin, MCH, and MCV, and normal to slightly decreased HbA2 levels.

• Alpha-Thalassemia Carriers (Alpha Thal Trait):

  • α+ trait (Silent Carrier): Asymptomatic, with normal RBC levels or slight microcytosis. Neonates may have a minor amount (1% to 3%) of Hb Bart’s.
  • α° trait (Alpha-Thalassemia 1 Trait): Usually presents with slight anemia, mildly reduced MCV and MCH, RBC microcytosis, hypochromia, and anisopoikilocytosis. Neonates may have moderate Hb Bart’s (3% to 8%) and some Heinz inclusion bodies.
  • Alpha-thalassemia carriers often have increased RBC counts, differentiating them from IDA. Diagnosis is often incidental during routine health checks or antenatal screening, highlighting the importance of recognizing alpha thal trait.

• Alpha-Thalassemia Minor: Results from two α-chain gene deletions. Patients have mild to moderate anemia (hemoglobin 7-10 g/dL). HbA2 levels are slightly elevated (3%–3.5%), and HbF levels may be slightly elevated (up to 5%). Molecular testing provides more definitive alpha thal trait diagnosis and differentiation.

• Alpha-Thalassemia Intermedia (HbH Disease): Characterized by anemia with varying HbH levels (0.8%–40%). Clinical severity varies by mutation type. Patients often present with severe microcytic or normocytic anemia early in life, with hemoglobin levels below 7 g/dL. Peripheral blood film shows poikilocytosis, tear-drop cells, increased erythroblasts, and target cells. HbH inclusion bodies are detectable with brilliant cresyl blue staining.

• Alpha-Thalassemia Major (Hb Bart’s Hydrops Fetalis Syndrome): Defined by the presence of Hb Bart’s and absence of HbF. Fetal blood HPLC shows sharp peaks of Hb Bart’s and Hb Portland. Alkaline electrophoresis shows Hb Bart’s in the anodal position and Hb Portland at the HbA position. Mothers may present with pregnancy-induced hypertension and polyhydramnios. Fetal ultrasound reveals hydrops. Cordocentesis shows severe fetal anemia (hemoglobin < 80 g/L). Laboratory investigation of parents shows decreased hemoglobin, MCH, and MCV, hypochromic, microcytic red cells on smear, and positive HbH inclusion body test, confirming carrier status and aiding in alpha thal trait diagnosis within the family.

Clinical Significance of Alpha Thal Trait Diagnosis

Accurate alpha thal trait diagnosis and diagnosis of all forms of α-thalassemia are critical for management decisions, such as whether to proceed with intrauterine transfusions for fetuses with Hb Bart’s hydrops fetalis. For parents with silent carrier or α-thalassemia minor traits, proper genetic counseling is essential. Molecular analysis is valuable in complex cases or families with mild mutations. Laboratory evaluation of α-thalassemia is paramount for improving patient quality of life through timely therapy, counseling, and complication prevention. Early and accurate alpha thal trait diagnosis in parents allows for informed reproductive choices and prenatal planning.

Quality Control and Lab Safety in Alpha-Thalassemia Testing

Laboratory quality control is essential for accurate alpha thal trait diagnosis and all thalassemia testing. Laboratories must establish internal quality control and participate in external quality assurance systems (EQAS) to ensure methodology precision and reliability. Standard operating procedures must be strictly followed. Reference standards and controls must be meticulously tracked. Levey Jennings charts can help monitor daily controls and identify errors.

A formal safety program is crucial for laboratory safety, including using protective gear (gloves, masks, eyewear, gowns) when handling specimens. Hand hygiene and safe disposal of sharps are essential. Adherence to safety protocols minimizes errors and exposure to hazardous materials, ensuring reliable alpha thal trait diagnosis and safe laboratory practices.

Enhancing Healthcare Team Outcomes in Alpha-Thalassemia Management

Effective diagnosis and management of α-thalassemia, including alpha thal trait, require a collaborative interprofessional healthcare team:

• Hematologist: Diagnoses and manages α-thalassemia, interprets lab results, and oversees treatment.
• Genetic Counselor: Provides genetic information and counseling, assesses transmission risks, explains test results, and assists with family planning related to alpha thal trait and other forms.
• Laboratory Technologist: Performs lab tests, ensures accuracy and timely reporting.
• Pediatrician/Internal Medicine Physician: Provides overall patient care.
• Obstetrician/Gynecologist: Manages high-risk pregnancies and fetal complications.
• Nurse Practitioner: Provides ongoing care, education, and support.
• Pharmacist: Manages medications, such as iron chelators.
• Transfusion Specialist: Manages blood transfusions.
• Clinical Geneticist: Consulted for complex genetic cases.

Effective communication and collaboration among these professionals are essential for comprehensive, patient-centered care, ensuring optimal outcomes for individuals and families affected by alpha-thalassemia, including those identified with alpha thal trait.

Review Questions

Figure

Hemoglobin Electrophoresis in Alpha-Thalassemia Major. This hemoglobin electrophoresis result demonstrates the presence of HbH and Hb Bart’s along with minimal HbA in a patient sample indicative of severe alpha-thalassemia.

Figure

Peripheral Blood Smear in Hydrops Fetalis Case. This peripheral blood film illustrates immature red cell precursors and red blood cells exhibiting hypochromia, microcytosis, anisocytosis, and poikilocytosis, characteristic of severe alpha-thalassemia leading to hydrops fetalis.

References

1.Thein SL. Molecular basis of β thalassemia and potential therapeutic targets. Blood Cells Mol Dis. 2018 May;70:54-65. [PMC free article: PMC5738298] [PubMed: 28651846]
2.Thein SL. The molecular basis of β-thalassemia. Cold Spring Harb Perspect Med. 2013 May 01;3(5):a011700. [PMC free article: PMC3633182] [PubMed: 23637309]
3.Bernini LF, Harteveld CL. Alpha-thalassaemia. Baillieres Clin Haematol. 1998 Mar;11(1):53-90. [PubMed: 10872473]
4.Harewood J, Azevedo AM. StatPearls [Internet]. StatPearls Publishing; Treasure Island (FL): Sep 4, 2023. Alpha Thalassemia. [PubMed: 28722856]
5.Weatherall DJ. The Evolving Spectrum of the Epidemiology of Thalassemia. Hematol Oncol Clin North Am. 2018 Apr;32(2):165-175. [PubMed: 29458724]
6.Munkongdee T, Chen P, Winichagoon P, Fucharoen S, Paiboonsukwong K. Update in Laboratory Diagnosis of Thalassemia. Front Mol Biosci. 2020;7:74. [PMC free article: PMC7326097] [PubMed: 32671092]
7.Rund D. Thalassemia 2016: Modern medicine battles an ancient disease. Am J Hematol. 2016 Jan;91(1):15-21. [PubMed: 26537527]
8.Musallam KM, Rivella S, Vichinsky E, Rachmilewitz EA. Non-transfusion-dependent thalassemias. Haematologica. 2013 Jun;98(6):833-44. [PMC free article: PMC3669437] [PubMed: 23729725]
9.Piel FB, Weatherall DJ. The α-thalassemias. N Engl J Med. 2014 Nov 13;371(20):1908-16. [PubMed: 25390741]
10.Poyart C, Wajcman H. Hemolytic anemias due to hemoglobinopathies. Mol Aspects Med. 1996 Apr;17(2):129-42. [PubMed: 8813715]
11.He S, Li J, Huang P, Zhang S, Lin L, Zuo Y, Tian X, Zheng C, Qiu X, Chen B. Characterization of Hb Bart’s Hydrops Fetalis Caused by – -SEA and a Large Novel α0-Thalassemia Deletion. Hemoglobin. 2018 Jan;42(1):61-64. [PubMed: 29493331]
12.Pang W, Long J, Weng X, Fan Q, Sun L, Pan Z, Fan Z. Identification of Three Types of α-Thalassemia Deletion, -α21.9, -α2.4, and – -THAI, and Their Frequencies, in One Family in the Population of Southern Guangxi Zhuang Autonomous Region, People’s Republic of China. Hemoglobin. 2018 Jan;42(1):37-42. [PubMed: 29447013]
13.Williams TN, Weatherall DJ. World distribution, population genetics, and health burden of the hemoglobinopathies. Cold Spring Harb Perspect Med. 2012 Sep 01;2(9):a011692. [PMC free article: PMC3426822] [PubMed: 22951448]
14.Kalfa TA. Diagnosis and clinical management of red cell membrane disorders. Hematology Am Soc Hematol Educ Program. 2021 Dec 10;2021(1):331-340. [PMC free article: PMC8791164] [PubMed: 34889366]
15.Bhat VS, Dewan KK, Krishnaswamy PR. The Diagnosis of α-Thalassaemia: A Case of Hemoglobin H -α Deletion. Indian J Clin Biochem. 2010 Oct;25(4):435-40. [PMC free article: PMC2994557] [PubMed: 21966120]
16.Viprakasit V, Limwongse C, Sukpanichnant S, Ruangvutilert P, Kanjanakorn C, Glomglao W, Sirikong M, Utto W, Tanphaichitr VS. Problems in determining thalassemia carrier status in a program for prevention and control of severe thalassemia syndromes: a lesson from Thailand. Clin Chem Lab Med. 2013 Aug;51(8):1605-14. [PubMed: 23525874]
17.King AJ, Higgs DR. Potential new approaches to the management of the Hb Bart’s hydrops fetalis syndrome: the most severe form of α-thalassemia. Hematology Am Soc Hematol Educ Program. 2018 Nov 30;2018(1):353-360. [PMC free article: PMC6246003] [PubMed: 30504332]
18.Giordano PC. Strategies for basic laboratory diagnostics of the hemoglobinopathies in multi-ethnic societies: interpretation of results and pitfalls. Int J Lab Hematol. 2013 Oct;35(5):465-79. [PubMed: 23217050]
19.Brancaleoni V, Di Pierro E, Motta I, Cappellini MD. Laboratory diagnosis of thalassemia. Int J Lab Hematol. 2016 May;38 Suppl 1:32-40. [PubMed: 27183541]
20.Hansson K, Schuring-Blom GH, Leschot NJ. The preserving of chorionic villi before establishing long-term cell cultures for cytogenetic analysis. Prenat Diagn. 1995 Nov;15(11):1067-9. [PubMed: 8606886]
21.Old J, Henderson S. Molecular diagnostics for haemoglobinopathies. Expert Opin Med Diagn. 2010 May;4(3):225-40. [PubMed: 23488532]
22.Galanello R, Origa R. Beta-thalassemia. Orphanet J Rare Dis. 2010 May 21;5:11. [PMC free article: PMC2893117] [PubMed: 20492708]
23.Oyaert M, Van Laer C, Claerhout H, Vermeersch P, Desmet K, Pauwels S, Kieffer D. Evaluation of the Sebia Minicap Flex Piercing capillary electrophoresis for hemoglobinopathy testing. Int J Lab Hematol. 2015 Jun;37(3):420-5. [PubMed: 25324031]
24.Chong SS, Boehm CD, Higgs DR, Cutting GR. Single-tube multiplex-PCR screen for common deletional determinants of alpha-thalassemia. Blood. 2000 Jan 01;95(1):360-2. [PubMed: 10607725]
25.Liu YT, Old JM, Miles K, Fisher CA, Weatherall DJ, Clegg JB. Rapid detection of alpha-thalassaemia deletions and alpha-globin gene triplication by multiplex polymerase chain reactions. Br J Haematol. 2000 Feb;108(2):295-9. [PubMed: 10691858]
26.Rao S, Kar R, Gupta SK, Chopra A, Saxena R. Spectrum of haemoglobinopathies diagnosed by cation exchange-HPLC & modulating effects of nutritional deficiency anaemias from north India. Indian J Med Res. 2010 Nov;132(5):513-9. [PMC free article: PMC3028946] [PubMed: 21150000]
27.Kunwandee J, Srivorakun H, Fucharoen G, Sanchaisuriya K, Fucharoen S. ARKRAY ADAMS A1c HA-8180T Analyzer for Diagnosis of Thalassemia and Hemoglobinopathies Common in Southeast Asia. Lab Med. 2014 Summer;45(3):e112-21. [PubMed: 25217515]
28.Khera R, Singh T, Khuana N, Gupta N, Dubey AP. HPLC in characterization of hemoglobin profile in thalassemia syndromes and hemoglobinopathies: a clinicohematological correlation. Indian J Hematol Blood Transfus. 2015 Mar;31(1):110-5. [PMC free article: PMC4275515] [PubMed: 25548455]
29.Borbely N, Phelan L, Szydlo R, Bain B. Capillary zone electrophoresis for haemoglobinopathy diagnosis. J Clin Pathol. 2013 Jan;66(1):29-39. [PubMed: 23105123]
30.Khongthai K, Ruengdit C, Panyasai S, Pornprasert S. Analysis of Deletional Hb H Diseases in Samples with Hb A2-Hb H and Hb A2-Hb Bart’s on Capillary Electrophoresis. Hemoglobin. 2019 Jul-Sep;43(4-5):245-248. [PubMed: 31687860]
31.Suwannakhon N, Pangeson T, Seeratanachot T, Mahingsa K, Pingyod A, Bumrungpakdee W, Sanguansermsri T. Noninvasive prenatal screening test for compound heterozygous beta thalassemia using an amplification refractory mutation system real-time polymerase chain reaction technique. Hematol Rep. 2019 Sep 18;11(3):8124. [PMC free article: PMC6761473] [PubMed: 31579144]
32.Qin J, He J, Li Y, Liu N, Tao F, Zhang P, Guo W, Qin Q, Zhou W. One-step genotyping of α-thalassaemia by multiplex symmetric PCR melting curve. J Clin Pathol. 2023 Sep;76(9):632-636. [PubMed: 35701141]
33.Green RC, Berg JS, Grody WW, Kalia SS, Korf BR, Martin CL, McGuire AL, Nussbaum RL, O’Daniel JM, Ormond KE, Rehm HL, Watson MS, Williams MS, Biesecker LG., American College of Medical Genetics and Genomics. ACMG recommendations for reporting of incidental findings in clinical exome and genome sequencing. Genet Med. 2013 Jul;15(7):565-74. [PMC free article: PMC3727274] [PubMed: 23788249]
34.Zhou YQ, Xiao GF, Li LY, Li WD, Liu ZY, Zhu LF, Mo QH, Qu XJ, Xu XM. Evaluation of clinical application of gap-PCR as a routine method for alpha-thalassemia carrier detection. Di Yi Jun Yi Da Xue Xue Bao. 2002 May;22(5):434-6. [PubMed: 12390707]
35.Schouten JP, McElgunn CJ, Waaijer R, Zwijnenburg D, Diepvens F, Pals G. Relative quantification of 40 nucleic acid sequences by multiplex ligation-dependent probe amplification. Nucleic Acids Res. 2002 Jun 15;30(12):e57. [PMC free article: PMC117299] [PubMed: 12060695]
36.Slatko BE, Gardner AF, Ausubel FM. Overview of Next-Generation Sequencing Technologies. Curr Protoc Mol Biol. 2018 Apr;122(1):e59. [PMC free article: PMC6020069] [PubMed: 29851291]
37.Emms A, Castleman J, Allen S, Williams D, Kinning E, Kilby M. Next Generation Sequencing after Invasive Prenatal Testing in Fetuses with Congenital Malformations: Prenatal or Neonatal Investigation. Genes (Basel). 2022 Aug 24;13(9) [PMC free article: PMC9498826] [PubMed: 36140685]
38.Kingchaiyaphum B, Sanchaisuriya K, Fucharoen G, Chaibunruang A, Hess SY, Hinnouho GM, Barffour MA, Wessells KR, Kounnavong S, Fucharoen S. Hemoglobins F, A2 , and E levels in Laotian children aged 6-23 months with Hb E disorders: Effect of age, sex, and thalassemia types. Int J Lab Hematol. 2020 Jun;42(3):277-283. [PMC free article: PMC7318314] [PubMed: 32048804]
39.Todd D, Lai MC, Beaven GH, Huehns ER. The abnormal haemoglobins in homozygous alpha-thalassaemia. Br J Haematol. 1970 Jul;19(1):27-31. [PubMed: 5465779]
40.Harteveld CL, Higgs DR. Alpha-thalassaemia. Orphanet J Rare Dis. 2010 May 28;5:13. [PMC free article: PMC2887799] [PubMed: 20507641]
41.Curran M, Mikhael M, Sun WD, Lim J, Leung A, Morchi G, Chmait RH. Perinatal Management of Bart’s Hemoglobinopathy: Paradoxical Effects of Intrauterine, Transplacental, and Partial Exchange Transfusions. AJP Rep. 2020 Jan;10(1):e11-e14. [PMC free article: PMC6984954] [PubMed: 31993246]
42.Songdej D, Babbs C, Higgs DR., BHFS International Consortium. An international registry of survivors with Hb Bart’s hydrops fetalis syndrome. Blood. 2017 Mar 09;129(10):1251-1259. [PMC free article: PMC5345731] [PubMed: 28057638]
43.Ghosh K, Colah R, Manglani M, Choudhry VP, Verma I, Madan N, Saxena R, Jain D, Marwaha N, Das R, Mohanty D, Choudhary R, Agarwal S, Ghosh M, Ross C. Guidelines for screening, diagnosis and management of hemoglobinopathies. Indian J Hum Genet. 2014 Apr;20(2):101-19. [PMC free article: PMC4228561] [PubMed: 25400338]
44.Torcharus K, Sriphaisal T, Krutvecho T, Ketupanya A, Vuthiwong C, Suwanasophon C, Noonai A. Prenatal diagnosis of Hb Bart’s hydrops fetalis by PCR technique: Pramongkutklao experience. Southeast Asian J Trop Med Public Health. 1995;26 Suppl 1:287-90. [PubMed: 8629126]
45.Dame C, Albers N, Hasan C, Bode U, Eigel A, Hansmann M, Brenner R, Bartmann P. Homozygous alpha-thalassaemia and hypospadias–common aetiology or incidental association? Long-term survival of Hb Bart’s hydrops syndrome leads to new aspects for counselling of alpha-thalassaemic traits. Eur J Pediatr. 1999 Mar;158(3):217-20. [PubMed: 10094442]
46.Fenta DA, Ali MM. Factors Affecting Quality of Laboratory Result During Ordering, Handling, and Testing of the Patient’s Specimen at Hawassa University College of Medicine and Health Science Comprehensive Specialized Hospital. J Multidiscip Healthc. 2020;13:809-821. [PMC free article: PMC7450655] [PubMed: 32922023]
47.Kinns H, Pitkin S, Housley D, Freedman DB. Internal quality control: best practice. J Clin Pathol. 2013 Dec;66(12):1027-32. [PubMed: 24072731]
48.Laudus N, Nijs L, Nauwelaers I, Dequeker EMC. The Significance of External Quality Assessment Schemes for Molecular Testing in Clinical Laboratories. Cancers (Basel). 2022 Jul 28;14(15) [PMC free article: PMC9367251] [PubMed: 35954349]
49.Skitek M, Martinello F, Jerin A. How to Really Understand and Improve the System of Internal Quality Control and External Quality Assessment in the Accreditation Process of the Medical Laboratory? EJIFCC. 2022 Apr;33(1):23-27. [PMC free article: PMC9092718] [PubMed: 35645692]
50.Lock RJ. My approach to internal quality control in a clinical immunology laboratory. J Clin Pathol. 2006 Jul;59(7):681-4. [PMC free article: PMC1860423] [PubMed: 16803947]
51.Petersen PH, Ricós C, Stöckl D, Libeer JC, Baadenhuijsen H, Fraser C, Thienpont L. Proposed guidelines for the internal quality control of analytical results in the medical laboratory. Eur J Clin Chem Clin Biochem. 1996 Dec;34(12):983-99. [PubMed: 8986407]
52.Abu-Siniyeh A, Al-Shehri SS. Safety in Medical Laboratories: Perception and Practice of University Students and Laboratory Workers. Appl Biosaf. 2021 Sep;26(Suppl 1):S34-S42. [PMC free article: PMC9134342] [PubMed: 36032652]
53.Bayot ML, Limaiem F. StatPearls [Internet]. StatPearls Publishing; Treasure Island (FL): Jan 30, 2023. Biosafety Guidelines. [PubMed: 30725895]
54.Cornish NE, Anderson NL, Arambula DG, Arduino MJ, Bryan A, Burton NC, Chen B, Dickson BA, Giri JG, Griffith NK, Pentella MA, Salerno RM, Sandhu P, Snyder JW, Tormey CA, Wagar EA, Weirich EG, Campbell S. Clinical Laboratory Biosafety Gaps: Lessons Learned from Past Outbreaks Reveal a Path to a Safer Future. Clin Microbiol Rev. 2021 Jun 16;34(3):e0012618. [PMC free article: PMC8262806] [PubMed: 34105993]