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Background Glycated hemoglobin, or hemoglobin A1c (HbA1c), serves as a crucial marker for diagnosing diabetes and monitoring its progression. We aimed to assess the interference posed by common Hb variants on popular HbA1c measurement systems. Methods A total of 63 variant and nonvariant samples with target values assigned by the International Federation of Clinical Chemistry and Laboratory Medicine (IFCC)-calibrated methods were included. We assessed 6 methods for measuring HbA1c in the presence of HbS, HbC, HbD, HbE, and fetal hemoglobin (HbF): 2 cation-exchange high-performance liquid chromatography (HPLC) methods (Bio-Rad D-100 and HLC-723 G8), a capillary electrophoresis (CE) method (Sebia Capillarys 3 TERA), an immunoassay (Roche c501), an enzyme assay system (Mindray BS-600M), and a boronate affinity method (Primus Premier Hb9210). Results The HbA1c results for nonvariant samples from the 6 methods were in good agreement with the IFCC-calibrated method results. The Bio-Rad D-100, Capillarys 3, Mindray BS-600M, Premier Hb9210, and Roche c501 showed no interference from HbS, HbC, HbD, and HbE. Clinically significant interference was observed for the HLC-723 G8 standard mode. Elevated HbF levels caused significant negative biases for all 6 methods, which increased with increasing HbF concentration. Conclusion Elevated levels of HbF can severely affect HbA1c measurements by borate affinity, immunoassays, and enzyme assays. [ABSTRACT FROM AUTHOR]

AbstractHemoglobin Tacoma is known to potentially interfere HbA1c assays. The variant is common in Finland with prevalence of up to 2% regionally and cases are also reported in areas that have attracted Finnish immigrants, especially in Sweden and North America. Here, we investigated the effect of Hb Tacoma on seven HbA1c methods. 20 non-variant and 20 Hb Tacoma samples were measured with Tina-quant Gen. 3 (immunoassay, considered as reference) and the following point of care instruments: Afinion 2, HbA1c 501 (both utilizing boronate affinity), QuikRead go, cobas b 101, DCA Atellica, and Standard F (all immunoassays). Repeatability was also assessed by measuring both non-variant and Hb Tacoma samples five times each at two different levels. For non-variant samples, the mean relative bias with all methods was < ±4%, whereas for Hb Tacoma samples Standard F had 38% mean relative bias. In absolute bias, the difference was 17 mmol/mol on average and constant through the measured range. For other methods the mean relative bias for Hb Tacoma samples was < ±6%. The repeatability with all methods was similar for non-variant and Hb Tacoma samples and at highest 4.1% (mean CV% of two levels). The observed interference by Standard F is likely due to two-antibody assay design as Hb Tacoma has been shown to result in conformational change. This interference is clinically significant and highlight the need for better controlling and better understanding hemoglobin variants in HbA1c testing. [ABSTRACT FROM AUTHOR]

We present 3 cases of discordant results from screening hemoglobin A1c (HbA1c) measured by ion-exchange high-performance liquid chromatography (HPLC) all due to various forms of interference and flagged by the instrument as "suspected hemoglobin E (HbE)." The first case was due to a rare hemoglobin variant, later confirmed to be hemoglobin Hoshida, the second due to "true" heterozygous HbE, and the third a result of analytical artifact causing splitting of the HbA1c peak without an underlying variant hemoglobin. We examine the similarities in these cases along with the laboratory work-up to classify each cause of interference to demonstrate the wide array of potential causes for the suspected HbE flag and why it warrants proper work-up. Because there is no standardized method of reporting out hemoglobin variant interference in HbA1c measurement, we discuss our laboratory's process of investigating discordant HbA1c measurements and reporting results in cases with variant interference as 1 possible model to follow, along with discussing the associated laboratory, ethical, and clinical considerations. We also examine the structure of hemoglobin Hoshida, HbE, and conduct a brief literature review of previous reports. [ABSTRACT FROM AUTHOR]

ABSTRACTObjectives: In clinical practice, the majority of α-thalassaemia cases arise from deletions of the α-globin genes. However, a subset of cases is attributed to rare haemoglobin variants, which can manifest with borderline or normal screening results, potentially leading to missed diagnoses in clinical practice.Methods: Blood samples were collected from family members and underwent haematological, DNA and RNA analysis.Results: The five-month-old proband presented a haematological phenotype consistent with Hb H disease. The mother’s haematology profile was consistent with an α-thalassaemia carrier, while the father exhibited a borderline reduction in MCV and MCH. MALDI-TOF identified an abnormal α-chain in the proband. DNA analysis revealed a novel α-globin variant (HBA2:c.175C>A, α58His>Asn, Hb DG-Nancheng) affecting the distal histidine in the family. The father and the mother had α-genotype of --SEA/αα and αDG-Nanchengα/αα, respectively; while the proband inherited both mutant alleles (--SEA/αDG-Nanchengα). Sequencing of cDNA from HBA2 gene identified an equal ratio of normal and mutant alleles.Conclusion: This rare case highlighted the importance of identifying rare haemoglobin variant during prenatal screening. The clinical and genetic data provides useful information on the pathogenicity of this variant and further insight into the role of distal histidine residue of α-globin.

Hemoglobin variants are often discovered when hemoglobin A1c (HbA1c) levels measured with a high-performance liquid chromatography (HPLC) system in fast mode are found to be low. The HA-8180V HPLC analyzer by Arkray offers two measurement modes: fast mode (FM) and variant mode (VM). Two Japanese patients with α-chain variant Hb Q-Iran detected incidentally after analyses with the HA-8180V in VM showed an abnormal peak, are presented. The first patient was a man in his 70 s, and the second patient was a man in his 50 s. Both were non-diabetic, but their results from HbA1c measurement in VM showed an abnormal peak. The VM–HbA1c, FM–HbA1c, and HbA1c measured by enzymatic assay and glycated albumin levels of the two patients were all within the reference ranges. They were diagnosed as having Hb Q-Iran (α2-75Asp → His) by globin gene analysis. It is difficult to detect α-chain hemoglobin variants based on abnormal FM–HbA1c levels, but measuring HbA1c in VM is useful for efficiently detecting hemoglobin variants. [ABSTRACT FROM AUTHOR]

Hemoglobin (Hb) Volga is a rare, unstable β-chain hemoglobin variant (β27 Ala→Asp), causing chronic hemolytic anemia. This study presents two members of a Danish family, splenectomized due to Hb Volga at and with multiple thrombotic events. The proband was diagnosed with Hb Volga 9 years old and splenectomy was performed as a part of treatment. Throughout his life, he experienced multiple superficial thrombophlebitis, two episodes of distal deep venous thrombosis (DVT) on lower extremities (age 32 and 33) and a transient ischemic attack (TIA) presented as amaurosis fugax (age 51). Thrombophilia investigation was normal. The proband's son was diagnosed with Hb Volga and underwent splenectomy at the age of 6. Despite anticoagulation therapy, he suffered from multiple venous thromboembolic events in his youth and died of chronic pulmonary embolism (PE)/pulmonary hypertension combined with infection. Given the observed propensity for multiple thromboses in these two patients, a literature review was conducted investigating reported occurrence of thrombotic events in individuals with Hb Volga. Currently 25 cases of Hb Volga are reported worldwide. The clinical symptoms primarily described are related to hemolytic anemia. Splenectomy is reported in 15 patients. Thromboses have previously been reported in only three patients who were also splenectomized. These cases involved DVT and PE, myocardial infarction, and an unspecified thrombotic event. The proband represents the first reported Hb Volga case with both venous and arterial thrombotic disorders. The exact mechanism underlying thrombotic tendency in patients with Hb Volga remains unknown, but it is probably associated with splenectomy. [ABSTRACT FROM AUTHOR]

Abstract Background Hemoglobin (Hb) Chile [β28(B10) Leu > Met; HBB: c.85 C > A] is a rare hemoglobin variant caused by a missense mutation in the HBB gene. Only one case of Hb Chile has been reported worldwide so far. It is an unstable hemoglobin, characterized by cyanosis associated with chronic methemoglobinemia and hemolytic anemia induced by sulfonamides or methylene blue. Case presentation A 9-year-3-month-old girl had mild anemia of unknown etiology for more than 6 years. She had a slight pallor without other symptoms or signs. The complete blood count revealed normocytic normochromic anemia with a sometimes-elevated reticulocyte count, and the bone marrow cytology showed marked erythroid hyperplasia, but the tests related to hemolysis were normal. Therefore, the whole exome sequencing was performed and showed a heterozygous mutation for HBB: c.85 C > A. With asymptomatic methemoglobinemia confirmed later, she was eventually diagnosed with Hb Chile. Conclusions This is the first report of Hb Chile in China and the second worldwide. This case shows that Hb Chile is clinically heterogeneous and difficult to diagnose and expands our understanding on the clinical and hematological traits of the disease.

Background: Hemoglobin (Hb) Chile [β28(B10) Leu > Met; HBB: c.85 C > A] is a rare hemoglobin variant caused by a missense mutation in the HBB gene. Only one case of Hb Chile has been reported worldwide so far. It is an unstable hemoglobin, characterized by cyanosis associated with chronic methemoglobinemia and hemolytic anemia induced by sulfonamides or methylene blue. Case presentation: A 9-year-3-month-old girl had mild anemia of unknown etiology for more than 6 years. She had a slight pallor without other symptoms or signs. The complete blood count revealed normocytic normochromic anemia with a sometimes-elevated reticulocyte count, and the bone marrow cytology showed marked erythroid hyperplasia, but the tests related to hemolysis were normal. Therefore, the whole exome sequencing was performed and showed a heterozygous mutation for HBB: c.85 C > A. With asymptomatic methemoglobinemia confirmed later, she was eventually diagnosed with Hb Chile. Conclusions: This is the first report of Hb Chile in China and the second worldwide. This case shows that Hb Chile is clinically heterogeneous and difficult to diagnose and expands our understanding on the clinical and hematological traits of the disease. [ABSTRACT FROM AUTHOR]

Introduction: Mutations in the hemoglobin subunit delta (HBD) gene (MIM#142000) are associated with decreased levels of the Hemoglobin A2 (Hb A2) fraction. We aimed to examine the prevalence of HBD gene mutations and summarize their characteristics in the Chinese population. Methods: Individuals who exhibited Hb A2 levels below 1.8%, with or without Hb A2 variant peaks, were chosen for further investigation. Hemoglobin analysis was conducted using capillary electrophoresis. Common α and β‐thalassemia in China were detected using gap‐PCR and reverse dot blot hybridization. The presence of HBD gene mutations was confirmed by DNA sequencing. Results: A total of 188 patients were identified as carriers of the HBD gene mutation, with a prevalence of approximately 0.46%. We discovered 36 types of mutations, 30 of which resulted in δ‐globin variants, while the remaining 6 resulted in δ‐thalassemia. The most common mutation was HBD:c.‐127 T > C, accounting for 87.2% of δ‐thalassemia cases. In addition, we identified 11 novel HBD gene mutations and found 10 cases compounded with other common thalassemias. Conclusion: We observed a high prevalence of HBD gene mutations in southern China. Our findings provide a genetic basis for screening for δ‐thalassemia and enrich the spectrum of HBD gene mutations. [ABSTRACT FROM AUTHOR]

Hb Chapel Hill [Alpha2 74(EF3) Asp > Gly] results from an GAC > GGC substitution at codon 74 of the HBA1 or HBA2 genes. Hb Chapel Hill has not been reported since 1986. A heterozygous mutation, HBA2: c.224A > G, was identified in the proband, her father and sister. We compared the haematological and clinical data of this family with the data reported in the limited number of individuals. Having excluded iron deficiency, the Hb Chapel Hill was asymptomatic in heterozygous state. The cases presented here characterize cases in new techniques including capillary electrophoresis (CE). Two aberrant peaks were identified by CE, a major peak migrating in the zone 7 that correspond to Hb Chapel Hill (αChapel Hill2β2) and a minor peak migrating in the zone 1 that correspond to Hb Chapel Hill2 (αChapel Hill2δ2). Focusing on the variant expression, the Hb Chapel Hill plus Hb A2 variant were around 18.9–20.6% of total Hb in three members. This data will be useful for providing up-to-date and high quality information on the Hb Chapel Hill. [ABSTRACT FROM AUTHOR]

In the present study, two unrelated cases of Hb Q-Thailand heterozygosity unlinked with the (–α4.2/) α+-thalassemia deletion allele were identified by long-read single molecule real-time (SMRT) sequencing in southern China. The aim of this study was to report the hematological and molecular features as well as diagnostic aspects of the rare manifestation. Hematological parameters and hemoglobin analysis results were recorded. A suspension array system for routine thalassemia genetic analysis and long-read SMRT sequencing were applied in parallel for thalassemia genotyping. Traditional methods, including Sanger sequencing, multiplex gap-polymerase chain reaction (gap-PCR) and multiplex ligation-dependent probe amplification (MLPA), were used together to confirm the thalassemia variants. Long-read SMRT sequencing was used to diagnose two Hb Q-Thailand heterozygous patients for whom the hemoglobin variant was unlinked to the (–α4.2/) allele for the first time. The hitherto undescribed genotypes were verified by traditional methods. Hematological parameters were compared with those of Hb Q-Thailand heterozygosity linked with the (–α4.2/) deletion allele in our study. For the positive control samples, long-read SMRT sequencing revealed a linkage relationship between the Hb Q-Thailand allele and the (–α4.2/) deletion allele. Identification of the two patients confirms that the linkage relationship between the Hb Q-Thailand allele and the (−α4.2/) deletion allele is a common possibility but not a certainty. Remarkably, as it is superior to traditional methods, SMRT technology may eventually serve as a more comprehensive and precise method that holds promising prospects in clinical practice, especially for rare variants. [ABSTRACT FROM AUTHOR]

Background: Hemoglobin A1c has been widely used to diagnose and monitor diabetes. However, the accuracy of HbA1c analysis can be significantly affected by hemoglobin variants, leading to falsely low or elevated levels and misdiagnosis or inappropriate diabetes management. Case report: In this study, we present the case of a 23-year-old man with undetectable HbA1c levels during his annual checkup by high-performance liquid chromatography (HPLC) and capillary electrophoresis (CE). To investigate the reason for HbA1c absence, Sanger sequencing, multiplex ligation-dependent probe amplification assay (MLPA), long-read single molecule real-time sequencing (SMRT) and MALDI-TOF mass spectrometry (MS) were performed, and the proband was identified as compound heterozygous of β-thalassemia with Hb G-Taipei (HBB:c.68A > G) and Hb Lepore-Boston-Washington (NG_000007.3:g.63632_71046del). Conclusion: The combination of these molecular technologies including MLPA, long-read SMRT sequencing and MALDI-TOF MS is beneficial for identifying rare hemoglobin variants. This case also provides essential evidence for uncovering the effect of compound heterozygosity for Hb Lepore-Boston-Washington and Hb G-Taipei on hematological phenotypes and HbA1c analysis.

Objectives: High-performance liquid chromatography (HPLC) is commonly used to measure hemoglobin A1c (HbA1c) levels and detect hemoglobin variants (Hb-Vars). HLC-723GR01 (GR01) is a new-generation automated ion-exchange HPLC system with two switchable analysis modes, namely short (30 s/test) and long modes (50 s/test). We evaluated the general performance of both analysis modes of GR01 for quantifying HbA1c and detecting Hb-Vars. Design and methods: We evaluated the instrument's precision based on CLSI protocol EP-05-A3. A comparison of the two analysis modes of GR01 against the standard mode of HLC-723G11 was performed on 100 whole blood samples. The GR01 long mode was compared with affinity HPLC (AF-HPLC) for detecting common Hb-Vars (HbE, HbD, HbS, and HbC, >20 samples). To examine the detection capability for minor Hb-Vars, we analyzed 26 Hb-Vars using multiple analyzers, including both analysis modes of GR01. Results: Both modes of GR01 had within-laboratory coefficients of variation of ≤1.0 % from four samples with HbA1c concentrations of 32–86 mmol/mol. Good correlation was observed between GR01 and HLC-723G11. The results for HbA1c detection in the presence of the major variants revealed a strong correlation between the long mode of GR01 and AF-HPLC (r = 0.986–0.998), and the difference biases ranged 0.1–1.9 mmol/mol. In the long mode, only one variant had a difference bias exceeding 14 % [10 % (%NGSP)]. Conclusion: The two analysis modes of GR01 were fast and had high accuracy and reproducibility, indicating their utility for routine clinical use in measuring HbA1c samples with Hb-Vars.

ABSTRACTBackground Hb Chapel Hill [Alpha2 74(EF3) Asp > Gly] results from an GAC > GGC substitution at codon 74 of the HBA1 or HBA2 genes. Hb Chapel Hill has not been reported since 1986.Methods A heterozygous mutation, HBA2: c.224A > G, was identified in the proband, her father and sister. We compared the haematological and clinical data of this family with the data reported in the limited number of individuals.Results Having excluded iron deficiency, the Hb Chapel Hill was asymptomatic in heterozygous state. The cases presented here characterize cases in new techniques including capillary electrophoresis (CE). Two aberrant peaks were identified by CE, a major peak migrating in the zone 7 that correspond to Hb Chapel Hill (αChapel Hill 2β2) and a minor peak migrating in the zone 1 that correspond to Hb Chapel Hill2 (αChapel Hill 2δ2). Focusing on the variant expression, the Hb Chapel Hill plus Hb A2 variant were around 18.9–20.6% of total Hb in three members.Conclusion This data will be useful for providing up-to-date and high quality information on the Hb Chapel Hill.

ABSTRACTObjective In the present study, two unrelated cases of Hb Q-Thailand heterozygosity unlinked with the (–α4.2/) α+-thalassemia deletion allele were identified by long-read single molecule real-time (SMRT) sequencing in southern China. The aim of this study was to report the hematological and molecular features as well as diagnostic aspects of the rare manifestation.Methods Hematological parameters and hemoglobin analysis results were recorded. A suspension array system for routine thalassemia genetic analysis and long-read SMRT sequencing were applied in parallel for thalassemia genotyping. Traditional methods, including Sanger sequencing, multiplex gap-polymerase chain reaction (gap-PCR) and multiplex ligation-dependent probe amplification (MLPA), were used together to confirm the thalassemia variants.Results Long-read SMRT sequencing was used to diagnose two Hb Q-Thailand heterozygous patients for whom the hemoglobin variant was unlinked to the (–α4.2/) allele for the first time. The hitherto undescribed genotypes were verified by traditional methods. Hematological parameters were compared with those of Hb Q-Thailand heterozygosity linked with the (–α4.2/) deletion allele in our study. For the positive control samples, long-read SMRT sequencing revealed a linkage relationship between the Hb Q-Thailand allele and the (–α4.2/) deletion allele.Conclusions Identification of the two patients confirms that the linkage relationship between the Hb Q-Thailand allele and the (−α4.2/) deletion allele is a common possibility but not a certainty. Remarkably, as it is superior to traditional methods, SMRT technology may eventually serve as a more comprehensive and precise method that holds promising prospects in clinical practice, especially for rare variants.

Keywords: hemoglobin variant; in-source decay; mass spectrometry; post-translational modification EN hemoglobin variant in-source decay mass spectrometry post-translational modification e251 e254 4 10/30/23 20231101 NES 231101 To the Editor, The diagnosis of hemoglobinopathy, routinely, involves a first-level analysis in which the peripheral hematological picture and the qualitative/quantitative profiling of the hemoglobins (Hbs) are assessed [[1]]. By allowing direct analysis of the hemolysate without the need for enzymatic digestion, the MALDI ISD analysis could represent a useful and rapid technique for the characterization of Hb variants, resulting also advantageous in the case of post-translational modified Hbs. [Extracted from the article]

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We present a new hemoglobin variant, Hb Raklev, characterized by the substitution of leucine with glutamine at position 75 in the β-globin chain. This variant was discovered inadvertently during an HbA1c evaluation using high performance liquid chromatography in a symptomless 54-year-old Caucasian woman, with the same variant also identified in her 16-year-old daughter. Purification of the hemoglobin revealed possibly diminished 2,3-bisphosphoglycerate (2,3-BPG) sensitivity, which may result in heightened oxygen affinity. Notably, two variants have been previously documented at this location: the unstable Hb Atlanta and the high-affinity Hb Pasadena. [ABSTRACT FROM AUTHOR]

Background and Aims: Mobility and migration flows are growing from different countries of the world to European countries, including France and in particular the Mediterranean basin. This study aimed to investigate the presence of hemoglobin (Hb) variants in outpatients/inpatients of the Montpellier Hospital (France) in whom an HbA1c assay had been performed and for which the country of birth had been informed. Methods: This is a retrospective study from January 2016 to December 2020 based on all high‐performance liquid chromatography (HPLC) chromatograms (Tosoh Bioscience HLC‐723G8) having an alarm of suspected Hb variant during HbA1c measurement. The corresponding samples were systematically sent to the hematology laboratory for confirmation and identification of Hb variant. Patient's medical history, clinical and demographic data were extracted from each medical chart. Statistical analyses were performed using XLSTAT® software, version 2016.06.35661. Results: Three hundred sixty‐three patients were confirmed with Hb variant exhibiting 17 different Hb profiles, highlighting the pivotal role of glycated hemoglobin (HbA1c) as a detection step. The prevalence of Hb variant in this southern French population was 0.71%, with the highest frequency for the beta‐globin variants (n = 342/363; i.e., 94.2%), including the most common: S, C, E, and D in 200/342 (58.5%), 83/342 (24.3%), 29/342 (8.5%), and 11/342 (3.2%), respectively. Among patients with Hb variants, almost half (165/363; i.e., 45.4%) were born in the African continent with a predominance for Morocco (32/165; i.e., 19.3%) and Algeria (29/165; i.e., 17.5%). Conclusion: HbA1c assay is a useful tool to detect Hb variants. Hemoglobinopathies are a public health issue in the current French population which is a multiethnic society. Despite the monocentric nature of our study, we note a high frequency of Hb variants in the south of France, which underlines the importance of screening for Hb variants in the whole population. [ABSTRACT FROM AUTHOR]

Objectives Here we report two rare α-globin chain variants in two unrelated families: Hb Val de Marne [α133(H16) Ser > Arg (AGC > CGC); HBA2: c.400A > C] and Hb Dongguan [α52(E6) Ser > Cys (TCT > TGT); HBA1: c.158C > G]. Notably, HBA2: c.400A > C is an unreported new variant in the third exon of the α2 gene, and simple heterozygous unstable Hb Dongguan haematological characteristics are proposed for the first time.Methods Hb analysis was performed by using capillary electrophoresis (CE). Twenty-three common mutations were detected using a suspension array system. Mutations were identified by DNA sequencing.Results The CE results showed an abnormal peak with incomplete separation from Hb A at zone 8 in two members of Family 1. DNA sequencing confirmed the presence of Hb Val de Marne [α133(H16) Ser > Arg (AGC > CGC); HBA2: c.400A > C]. Five members of Family 2 exhibited an abnormal peak at zone 11, and DNA sequencing confirmed the presence of Hb Dongguan [α52(E6) Ser > Cys (TCT > TGT); HBA1: c.158C > G].Conclusions The discovery of HBA2: C.400A > C expands the existing spectrum of α-globin variants. The carriers of simple heterozygous Hb Dongguan generally do not have obvious clinical symptoms. The information in this study will help clinicians understand the screening, molecular diagnosis and clinical significance of Hb variants.

Objectives Shaokwan is one of the inhabitant regions of Hakka population in Guangdong province of southern China. Previous survey has reported that a higher prevalence of abnormal hemoglobin (Hb) in this region. However, large-scale survey on the molecular characteristics of abnormal hemoglobin in this south-north junctional region has not been reported.In this study, we aim to investigate the molecular characteristics of abnormal hemoglobin in this area. Methods Blood samples from medical check-up center in one local hospital were selected for abnormal hemoglobin screening. Hemoglobin electrophoresis and routine blood tests were performed. Hemoglobin variants were further analyzed by PCR and DNA sequencing. Results Among the 9731 study subjects, 45 cases of hemoglobin variants were found by hemoglobin electrophoresis, which gave an incidence of 0.46% (45/9731) in Shaokwan region. 8 kinds of hemoglobin variants were identified by gene sequencing. Hb Q-Thailand (16/45) was the most common hemoglobin variants, followed by HbE (7/45), Hb NewYork (6/45) and Hb G-Chinese (6/45). Discussion and Conclusion Hemoglobin variants had obviously genetic polymorphisms in Chinese. Our study of hemoglobin disorders in this special Hakka Chinese population will contribute considerably to our understanding of the historical, emigrational, and genetic relationships among different ethnic group in this region.

Hemoglobin Santa Ana [β88(F4)Leu→Pro (CTG > CCG) HBB: c.266T > C] is an unstable hemoglobin variant characterized by a substitution of the amino acid leucine by proline at the 88th position of the β-globin chain. We for the first time identified this hemoglobin variant in a Chinese patient by capillary electrophoresis (CE). The proband was an 8-year-old boy with chronic anemia, brown urine and splenomegaly. He had been affected by moderate anemia, twice approaching a severe degree, that was attributed to infection. The CE result revealed an abnormal hemoglobin peak at electrophoretic zone 4 that correspond to the hemoglobin Santa Ana peak, and a CTG > CCG mutation at codon 88 of the β-globin gene was confirmed by DNA sequencing. To avoid misdiagnosis and genetic risks, a literature review of other unstable hemoglobins that migrate similarly to the hemoglobin Santa Ana was performed. Our findings indicate that hemoglobin Santa Ana can be clearly separated by CE, with accurate diagnosis depending on molecular analysis. This information will be useful for providing appropriate genetic counselling and for prenatal diagnosis.

Background HbA1c is the validated biomarker for glycemic management in diabetic individuals. Here, we report a compound heterozygote for β0-thal and Hb J-Lomeand evaluate its effect on HbA1c measurements.Methods A 51-year-old female was suspected of harboring a hemoglobin variant following no value of HbA1c levelby Arkray HA-8180 V (48s HbA1c mode), abnormal hematological data, and abnormalhemoglobin analysison capillary electrophoresis (Capillarys 2 Flex Piercing, Hb program). Sanger sequencing of the α and β genes was subsequently performed on the proband.HbA1c was reanalyzed using D10 (Bio-Rad), Capillarys 2 Flex Piercing (Sebia), and Roche Cobas c501 (Roche Diagnostics).Results Sanger sequencing identified a compound heterozygote for β0-thal [β17(A14) Lys > Stop, HBB: c.52A > T] and Hb J-Lome [β59(E3) Lys > Asn, HBB: c.180G > C].HbA1c values ⁣⁣determinedby D10, Capillarys 2 Flex Piercing (HbA1c program), and Roche Cobas c501were 2.3%, no HbA1c value, and 5.1 (32 mmol/mol), respectively. During pedigree analysis, the son of the proband was found to have normal blood glucose (5.55 mmol/L), decreased HbA1c (3.6%, 16 mmol/mol)by Arkray HA-8180 V (48s HbA1c mode), an abnormal band on the electrophoretogram of Capillarys2 (Hb program), and the Hb J-Lome mutation in the β globin gene.Subsequently, HbA1c values ⁣⁣determinedby D10, Capillarys 2 Flex Piercing (HbA1c program), and Roche Cobas c501 were4.0% (20 mmol/mol), no HbA1c value, and 5.0 (31 mmol/mol), respectively.Conclusion Atypically low HbA1c levels or a discrepancy between blood glucose and HbA1c levels should raise concerns about hemoglobin variations.

Abstract Background and Aims Mobility and migration flows are growing from different countries of the world to European countries, including France and in particular the Mediterranean basin. This study aimed to investigate the presence of hemoglobin (Hb) variants in outpatients/inpatients of the Montpellier Hospital (France) in whom an HbA1c assay had been performed and for which the country of birth had been informed. Methods This is a retrospective study from January 2016 to December 2020 based on all high‐performance liquid chromatography (HPLC) chromatograms (Tosoh Bioscience HLC‐723G8) having an alarm of suspected Hb variant during HbA1c measurement. The corresponding samples were systematically sent to the hematology laboratory for confirmation and identification of Hb variant. Patient's medical history, clinical and demographic data were extracted from each medical chart. Statistical analyses were performed using XLSTAT® software, version 2016.06.35661. Results Three hundred sixty‐three patients were confirmed with Hb variant exhibiting 17 different Hb profiles, highlighting the pivotal role of glycated hemoglobin (HbA1c) as a detection step. The prevalence of Hb variant in this southern French population was 0.71%, with the highest frequency for the beta‐globin variants (n = 342/363; i.e., 94.2%), including the most common: S, C, E, and D in 200/342 (58.5%), 83/342 (24.3%), 29/342 (8.5%), and 11/342 (3.2%), respectively. Among patients with Hb variants, almost half (165/363; i.e., 45.4%) were born in the African continent with a predominance for Morocco (32/165; i.e., 19.3%) and Algeria (29/165; i.e., 17.5%). Conclusion HbA1c assay is a useful tool to detect Hb variants. Hemoglobinopathies are a public health issue in the current French population which is a multiethnic society. Despite the monocentric nature of our study, we note a high frequency of Hb variants in the south of France, which underlines the importance of screening for Hb variants in the whole population.

Previous studies have identified occasional cases of heterozygous Hb Tacoma in areas that have attracted Finnish immigrants, especially in Sweden and North America, but large studies of this slightly unstable beta variant in vitro have not been carried out. Here we determined the prevalence of hemoglobin variants across Finland. A total of 5059 samples from 11 different hospital districts were analyzed using HbA1c capillary electrophoresis and reviewed for atypical profiles (HbA1c, Capillarys 3 Tera, Sebia). 38 heterozygous Hb Tacoma cases were found (0.75%). The prevalence was highest in South Ostrobothnia (2.0%), located in western Finland, and second highest in the neighboring provinces (1.0–1.4%), but only two districts were devoid of variants. Heterozygous Hb Tacoma was confirmed by genetic testing (NM_000518.5(HBB):c.93G > T (p.Arg31Ser)). In addition, five other variants were found, suggestive of HbE, Hb Helsinki (two cases) and an alpha variant, as interpreted from the electropherograms. The fifth variant, belonging to the South Ostrobothnian cohort, remained unknown at the time of the initial analyses, but was later interpreted as homozygous Hb Tacoma and confirmed by hemoglobin fraction analysis (Hemoglobin(E), Capillarys 3 Tera). In a subsequent retrospective study of the electropherograms of routine HbA1c diagnostics, altogether nine homozygous Hb Tacoma cases were identified in South Ostrobothnia. While heterozygous Hb Tacoma is usually an incidental finding, it interferes with several HbA1c assays. The present study is the first demonstration of homozygous Hb Tacoma. The clinical presentations of homozygous Hb Tacoma are not known and need to be addressed in future studies. [ABSTRACT FROM AUTHOR]

Hb variants prevalent in China are different from those in other countries. We aimed to assess the interference from Hb variants found in China on HbA1c measurement. All Hb variants were confirmed using Sanger sequencing. HbA1c was measured using a capillary electrophoresis method (Capillarys 3 OCTA), two cation-exchange high-performance liquid chromatography methods (ADAMS HA-8180V and HLC-723 G8 standard mode), an immunoassay (Cobas c501), and a boronate affinity chromatography method (Premier Hb9210). Premier Hb9210 was used as a comparative method. A total of 16 species of Hb variants were identified in 102 variant carriers. The most common variant was Hb E, followed by Hb Q-Thailand, Hb New York and Hb J-Bangkok. Clinically significant interference was observed for the Capillarys 3 OCTA (two Hb variants), ADAMS HA-8180V (seven Hb variants), HLC-723 G8 (14 Hb variants), and Cobas c501 (two Hb variants). The proportion of unacceptable HbA1c results was 13.7% for Capillarys 3 OCTA, 52.9% for HA-8180V, 83.3% for HLC-723 G8, and 3.9% for Cobas c501. Hb variants in China severely affect the accuracy of some commonly used HbA1c methods. [ABSTRACT FROM AUTHOR]

The patient, a man in his thirties, presented to our hospital for a secondary examination after a 2020 medical check-up found a high hemoglobin A1c (HbA1c) level on high-performance liquid chromatography (HPLC). The HbA1c level determined by HPLC (HA-8180V, fast mode) was elevated at 6.8%, but a 75-g glucose tolerance test showed normal glucose tolerance. The glycated albumin level was within the reference range at 14.6%. The continuous glucose monitoring-derived mean blood glucose and the percentage of time in range were 99 mg/dL and 98%, respectively. The HbA1c levels determined by HPLC (G9, fast mode), enzymatic assay, and immunoassay were all 5.3%. An isoelectric focusing analysis showed an abnormal band on the anode side of HbA2, and a globin gene analysis detected a heterozygous mutation at codon 144 [AAG (Lys) → TAG (stop codon)] in the δ-chain. Since this mutation is a novel δ-chain hemoglobin variant, it was given the name 'Hb A2-Karatsu'. [ABSTRACT FROM AUTHOR]

HbA1c is the validated biomarker for glycemic management in diabetic individuals. Here, we report a compound heterozygote for β0-thal and Hb J-Lomeand evaluate its effect on HbA1c measurements. A 51-year-old female was suspected of harboring a hemoglobin variant following no value of HbA1c levelby Arkray HA-8180 V (48s HbA1c mode), abnormal hematological data, and abnormalhemoglobin analysison capillary electrophoresis (Capillarys 2 Flex Piercing, Hb program). Sanger sequencing of the α and β genes was subsequently performed on the proband.HbA1c was reanalyzed using D10 (Bio-Rad), Capillarys 2 Flex Piercing (Sebia), and Roche Cobas c501 (Roche Diagnostics). Sanger sequencing identified a compound heterozygote for β0-thal [β17(A14) Lys > Stop, HBB: c.52A > T] and Hb J-Lome [β59(E3) Lys > Asn, HBB: c.180G > C].HbA1c values ⁣⁣determinedby D10, Capillarys 2 Flex Piercing (HbA1c program), and Roche Cobas c501were 2.3%, no HbA1c value, and 5.1 (32 mmol/mol), respectively. During pedigree analysis, the son of the proband was found to have normal blood glucose (5.55 mmol/L), decreased HbA1c (3.6%, 16 mmol/mol)by Arkray HA-8180 V (48s HbA1c mode), an abnormal band on the electrophoretogram of Capillarys2 (Hb program), and the Hb J-Lome mutation in the β globin gene.Subsequently, HbA1c values ⁣⁣determinedby D10, Capillarys 2 Flex Piercing (HbA1c program), and Roche Cobas c501 were4.0% (20 mmol/mol), no HbA1c value, and 5.0 (31 mmol/mol), respectively. Atypically low HbA1c levels or a discrepancy between blood glucose and HbA1c levels should raise concerns about hemoglobin variations. [ABSTRACT FROM AUTHOR]

Here we report two rare α-globin chain variants in two unrelated families: Hb Val de Marne [α133(H16) Ser > Arg (AGC > CGC); HBA2: c.400A > C] and Hb Dongguan [α52(E6) Ser > Cys (TCT > TGT); HBA1: c.158C > G]. Notably, HBA2: c.400A > C is an unreported new variant in the third exon of the α2 gene, and simple heterozygous unstable Hb Dongguan haematological characteristics are proposed for the first time. Hb analysis was performed by using capillary electrophoresis (CE). Twenty-three common mutations were detected using a suspension array system. Mutations were identified by DNA sequencing. The CE results showed an abnormal peak with incomplete separation from Hb A at zone 8 in two members of Family 1. DNA sequencing confirmed the presence of Hb Val de Marne [α133(H16) Ser > Arg (AGC > CGC); HBA2: c.400A > C]. Five members of Family 2 exhibited an abnormal peak at zone 11, and DNA sequencing confirmed the presence of Hb Dongguan [α52(E6) Ser > Cys (TCT > TGT); HBA1: c.158C > G]. The discovery of HBA2: C.400A > C expands the existing spectrum of α-globin variants. The carriers of simple heterozygous Hb Dongguan generally do not have obvious clinical symptoms. The information in this study will help clinicians understand the screening, molecular diagnosis and clinical significance of Hb variants. [ABSTRACT FROM AUTHOR]

Hemoglobin Santa Ana [β88(F4)Leu→Pro (CTG > CCG) HBB: c.266T > C] is an unstable hemoglobin variant characterized by a substitution of the amino acid leucine by proline at the 88th position of the β-globin chain. We for the first time identified this hemoglobin variant in a Chinese patient by capillary electrophoresis (CE). The proband was an 8-year-old boy with chronic anemia, brown urine and splenomegaly. He had been affected by moderate anemia, twice approaching a severe degree, that was attributed to infection. The CE result revealed an abnormal hemoglobin peak at electrophoretic zone 4 that correspond to the hemoglobin Santa Ana peak, and a CTG > CCG mutation at codon 88 of the β-globin gene was confirmed by DNA sequencing. To avoid misdiagnosis and genetic risks, a literature review of other unstable hemoglobins that migrate similarly to the hemoglobin Santa Ana was performed. Our findings indicate that hemoglobin Santa Ana can be clearly separated by CE, with accurate diagnosis depending on molecular analysis. This information will be useful for providing appropriate genetic counselling and for prenatal diagnosis. [ABSTRACT FROM AUTHOR]

Shaokwan is one of the inhabitant regions of Hakka population in Guangdong province of southern China. Previous survey has reported that a higher prevalence of abnormal hemoglobin (Hb) in this region. However, large-scale survey on the molecular characteristics of abnormal hemoglobin in this south-north junctional region has not been reported.In this study, we aim to investigate the molecular characteristics of abnormal hemoglobin in this area. Blood samples from medical check-up center in one local hospital were selected for abnormal hemoglobin screening. Hemoglobin electrophoresis and routine blood tests were performed. Hemoglobin variants were further analyzed by PCR and DNA sequencing. Among the 9731 study subjects, 45 cases of hemoglobin variants were found by hemoglobin electrophoresis, which gave an incidence of 0.46% (45/9731) in Shaokwan region. 8 kinds of hemoglobin variants were identified by gene sequencing. Hb Q-Thailand (16/45) was the most common hemoglobin variants, followed by HbE (7/45), Hb NewYork (6/45) and Hb G-Chinese (6/45). Hemoglobin variants had obviously genetic polymorphisms in Chinese. Our study of hemoglobin disorders in this special Hakka Chinese population will contribute considerably to our understanding of the historical, emigrational, and genetic relationships among different ethnic group in this region. [ABSTRACT FROM AUTHOR]

The case of a 49-year-old man with Hb Phnom Penh whose HbA1c levels varied depending on the high-performance liquid chromatography (HPLC) system is presented. After the results of a health checkup showed a high HbA1c level of 7.6% measured by HPLC method, another HbA1c test measured by immunoassay at a local clinic showed a level of 5.2%. Given the discrepancy in HbA1c levels between assay methods, he visited our clinic. His fasting plasma glucose level was 95 mg/dL, but an oral glucose tolerance test showed diabetes mellitus. The mean plasma glucose level was 117 mg/dL on intermittently scanned continuous glucose monitoring (isCGM), giving an estimated HbA1c level of 5.9%. Based on the results of isCGM, the present case was considered to be not diabetes mellitus. The same specimen was assayed for HbA1c by various methods (systems), including HPLC with Tosoh G9, HPLC with Arkray HA-8180 and HA-8180T, affinity assay, and enzymatic assay, giving HbA1c levels of 7.3%, 4.9%, 5.0%, 5.4%, and 5.3%, respectively. A globin gene analysis diagnosed Hb Phnom Penh ( α 1-117Phe-Ile- α 1-118Thr). The present case had a unique hemoglobin variant, resulting in high HbA1c levels when measured with Tosoh's HPLC systems and low HbA1c levels with Arkray's HPLC systems., (© 2024 by the Association of Clinical Scientists, Inc.)

Comparison of the Sebia 3-OCTA vs. the Tosoh-G8 showed a strong correlation (r=0.995), with a bias of -0.1% (-1.4 mmol/mol) (Supplemental Material, Figure A). Keywords: analytical interferences; comparison method; hemoglobin variant; imprecision EN analytical interferences comparison method hemoglobin variant imprecision e216 e220 5 08/05/22 20220801 NES 220801 To the Editor, Great effort has been made to standardize the methods for the measurement of HbA SB 1c sb , however, analytical interferences persist with some HbA SB 1c sb methods in presence of hemoglobin (Hb) variants, high levels of HbF or uremia. Special attention should be given to the results of HbA SB 1c sb in the presence of HbA/G-San Jose on Sebia 3-OCTA and the presence of HbA/E and HbA/D-Punjab on the Tosoh G8. Comparison of Sebia Capillarys 3-OCTA with the Tosoh Bioscience HLC®-723G8 method for A1C testing with focus on analytical interferences and variant detection. [Extracted from the article]

Objectives: We aimed to evaluate the effects of hemoglobin (Hb) variants prevalent in China on HbA1c measurements and to identify them during HbA1c measurements.Methods: We evaluated a cation-exchange high-performance liquid chromatography (HPLC) method (Bio-Rad D-100), a capillary electrophoresis (CE) method (Capillarys 3 TERA), an immunoassay (Cobas c501), and a boronate affinity method (Premier Hb9210, as a comparative method) for HbA1c measurements in the presence of Hb variants prevalent in China.Results: The Bio-Rad D-100 and Capillarys 3 TERA gave specific retention times and numeric migration positions for each Hb variant, respectively, showing excellent interindividual reproducibility. All methods showed statistically significant differences (P < .01) for several variants. Clinically significant effects were observed for the Bio-Rad D-100 (Hb New York and Hb J-Bangkok), Capillarys 3 TERA (Hb New York and Hb J-Bangkok), and Cobas c501 (Hb New York). Among 297 samples with Hb variants, there were 75 (25.3%) unacceptable results for Bio-Rad D-100, 28 (9.4%) for Capillarys 3 TERA, and 19 (6.4%) for Cobas c501 compared with the results from Premier Hb9210.Conclusions: Some Hb variants prevalent in China affect HbA1c measurements. The HPLC retention time and CE migration position can aid in the presumptive identification of Hb variants. [ABSTRACT FROM AUTHOR]

Background: Glycated Hemoglobin A1c (HbA1c) assay is most widely used in diabetic patients for assessing long-term control of glycemia. The presence of hemoglobin variants may be an incidental finding and can interfere with HbA1c measurements. The aim of the present study is to investigate the prevalence and impact of interference of different abnormal hemoglobin variants on HbA1c measurements during routine HbA1c testing. Methods: A total of 12,092 HbA1c samples were collected from January to August 2018. HbA1c quantification was carried out on a Variant II Bio-Rad’s HPLC analyzer. Abnormal chromatograms were further analyzed using the extended-run high-pressure liquid chromatography (HPLC) analysis in the A2/F mode. Results: The samples were examined for presence of abnormal variants. Samples producing abnormal chromatograms were further analyzed in A2/F mode to characterize hemoglobin variants. Abnormal variants were identified in 126 (1%) samples, and 74 (0.59%) sickle cell traits (SCT) were the most common variant in our findings. Moreover, 30 (0.24%) cases were eluted in the variant window in A1c mode, which on further analysis were found to be Hb E & Hb D traits. Furthermore, 3 (0.02%) cases were eluted at a RT

Background: High performance liquid chromatography (HPLC) is the most commonly used method for detection and quantitative estimation of hemoglobin variants. Hemoglobinopathies are amongst the most common genetically inherited disorders, however, the exact magnitude of different hemoglobinopathies is obscure in India. This study was done with the aim of analyzing the different findings in HPLC using D-10 analyzer and evaluating the spectrum of different hemoglobin disorders in a hospital-based population of South Delhi. Such a prevalence study would be useful to review the various strategies that can be implemented for effective control and prevention of these disorders. Methods: A hospital based descriptive observational study was conducted in which all OPD and IPD patients who were advised HPLC during their clinical workup were included. Analysis of EDTA blood samples was done by Bio Rad D10 Dual program HPLC instrument. The exact percentage of HbA, HbA2, HbF and any other variant hemoglobin was estimated. Presumptive identification of hemoglobin variants was made primarily by their percentage, retention time (RT) and peak characteristics. HPLC findings were correlated with the clinical history, family history and the CBC and peripheral smear findings in all cases. Results: On HPLC analysis, 79% of the patients had no abnormality detected and the report was within normal limits. The commonest hemoglobinopathy was Beta Thalassemia Trait followed by HbE trait. The other hemoglobinopathies detected were HbD Punjab Heterozygous (3 cases, 0.5%), Beta thalassemia homozygous (3 cases, 0.5%), Sickle cell Heterozygous (2 cases, 0.3%), HbJ Meerut Heterozygous (2 cases, 0.3%). One case each of Sickle cell Homozygous (0.15%), Compound Heterozygous HbS/beta thalassemia trait (0.15%), HbE Homozygous (0.15%), Compound Heterozygous HbE/beta thalassemia trait (0.15%), and Homozygous delta beta thalassemia (0.15%) were also diagnosed. Conclusion: This study gives an important insight to the present day scenario of hemoglobinopathies in patients in South Delhi in relation to the hematological profile. It highlights the chromatogram findings of different hemoglobinopathies on the D10 analyzer. The comprehensive data obtained by such series can help in the formulation and development of infrastructure and policies for hemoglobinopathy prevention, diagnosis and management.

Background: High performance liquid chromatography (HPLC) is the most commonly used method for detection and quantitative estimation of hemoglobin variants. Hemoglobinopathies are amongst the most common genetically inherited disorders, however, the exact magnitude of different hemoglobinopathies is obscure in India. This study was done with the aim of analyzing the different findings in HPLC using D-10 analyzer and evaluating the spectrum of different hemoglobin disorders in a hospital-based population of South Delhi. Such a prevalence study would be useful to review the various strategies that can be implemented for effective control and prevention of these disorders. Methods: A hospital based descriptive observational study was conducted in which all OPD and IPD patients who were advised HPLC during their clinical workup were included. Analysis of EDTA blood samples was done by Bio Rad D10 Dual program HPLC instrument. The exact percentage of HbA, HbA2, HbF and any other variant hemoglobin was estimated. Presumptive identification of hemoglobin variants was made primarily by their percentage, retention time (RT) and peak characteristics. HPLC findings were correlated with the clinical history, family history and the CBC and peripheral smear findings in all cases. Results: On HPLC analysis, 79% of the patients had no abnormality detected and the report was within normal limits. The commonest hemoglobinopathy was Beta Thalassemia Trait followed by HbE trait. The other hemoglobinopathies detected were HbD Punjab Heterozygous (3 cases, 0.5%), Beta thalassemia homozygous (3 cases, 0.5%), Sickle cell Heterozygous (2 cases, 0.3%), HbJ Meerut Heterozygous (2 cases, 0.3%). One case each of Sickle cell Homozygous (0.15%), Compound Heterozygous HbS/beta thalassemia trait (0.15%), HbE Homozygous (0.15%), Compound Heterozygous HbE/beta thalassemia trait (0.15%), and Homozygous delta beta thalassemia (0.15%) were also diagnosed. Conclusion: This study gives an important insight to the present day scenario of hemoglobinopathies in patients in South Delhi in relation to the hematological profile. It highlights the chromatogram findings of different hemoglobinopathies on the D10 analyzer. The comprehensive data obtained by such series can help in the formulation and development of infrastructure and policies for hemoglobinopathy prevention, diagnosis and management. [ABSTRACT FROM AUTHOR]

Glycated hemoglobin A1c (HbA1c) is considered the standard of care for the testing and monitoring of diabetes. Its ability to accurately reflect glycemia, however, is imperfect. Hemoglobin variants—mutant forms of hemoglobin caused by genetic variation present in 7% of the population—are known to adversely affect the ability of HbA1c measurement to reflect glycemic control. We report an illustrative case of a 64-year-old nondiabetic man with a steadily decreasing HbA1c and no symptoms of hypoglycemia or concerning family history. Preliminary investigative workup returned nothing of significance. Genetic sequencing, however, identified a rare benign hemoglobin variant: a heterozygous missense mutation in the gene encoding the hemoglobin β chain (c.155C > A, p.Pro51His). This variant has been reported only once previously, and the report predates genetic sequence data of the variant. Although this variant had no clinical implications for the patient, it was the cause of falsely low HbA1c levels on high-performance ion-exchange chromatography. This case highlights the importance of considering the effect of hemoglobin variants on the measurement of HbA1c. When available, family history should be carefully considered. Clinicians should suspect hemoglobin variants when HbA1c is too high or low, or discordant with the clinical picture. [ABSTRACT FROM AUTHOR]

Objectives: In clinical practice, the majority of α-thalassaemia cases arise from deletions of the α-globin genes. However, a subset of cases is attributed to rare haemoglobin variants, which can manifest with borderline or normal screening results, potentially leading to missed diagnoses in clinical practice., Methods: Blood samples were collected from family members and underwent haematological, DNA and RNA analysis., Results: The five-month-old proband presented a haematological phenotype consistent with Hb H disease. The mother's haematology profile was consistent with an α-thalassaemia carrier, while the father exhibited a borderline reduction in MCV and MCH. MALDI-TOF identified an abnormal α-chain in the proband. DNA analysis revealed a novel α-globin variant ( HBA2 :c.175C>A, α58His>Asn, Hb DG-Nancheng) affecting the distal histidine in the family. The father and the mother had α-genotype of -- SEA /αα and α DG-Nancheng α/αα, respectively; while the proband inherited both mutant alleles (-- SEA /α DG-Nancheng α). Sequencing of cDNA from HBA2 gene identified an equal ratio of normal and mutant alleles., Conclusion: This rare case highlighted the importance of identifying rare haemoglobin variant during prenatal screening. The clinical and genetic data provides useful information on the pathogenicity of this variant and further insight into the role of distal histidine residue of α-globin.

Abstract Hemoglobin (Hb) North Manchester [β51(D2) Pro→His; HBB:c.155 C>A] is a rare Hb β‐globin gene variant that affects glycated Hb measurement values, such as ion‐exchange high‐performance liquid chromatography, in patients with diabetes. This variant was first detected in the UK in 1998. Here, we describe the first case involving Hb North Manchester detected incidentally in a patient with type 2 diabetes in Northern China. The Hb variant was discovered by ion‐exchange high‐performance liquid chromatography, yet capillary electrophoresis of both glycated Hb program and Hb program failed to detect it. Subsequently, Sanger sequencing was carried out to help identify the Hb variant.

Objectives: The present study was conducted to evaluate the usefulness of capillary electrophoresis (CE), cation-exchange high-performance liquid chromatography (HPLC), and matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) for the screening of hemoglobin (Hb) variants prevalent in southern China.Methods: A total of 102 types of Hb variants in 1,083 variant carriers were identified over a 5-year period. These variants were analyzed by a CE method (Capillarys 3 TERA), a cation-exchange HPLC analyzer (Variant II Turbo 2.0), and a MALDI-TOF MS system (QuanTOF).Results: The presence of 85 (83.3%, 85/102), 84 (82.4%, 84/102), and 62 (60.8%, 62/102) Hb variants was detected by Capillarys 3 TERA, Variant II Turbo 2.0, and QuanTOF, respectively. Of the three methods, only Capillarys 3 TERA recognized all 10 of the most frequent Hb variants in southern China. There were six, two, and three Hb variants that can only be detected by Capillarys 3 TERA, Variant II Turbo 2.0, and QuanTOF, respectively. The detection limit of mass difference for QuanTOF was approximately 11 to 20 Da.Conclusions: MALDI-TOF MS is suitable for use as an auxiliary method rather than a stand-alone method for the screening of Hb variants prevalent in southern China. [ABSTRACT FROM AUTHOR]

Hemoglobin (Hb) variants, characterized by structural abnormalities in the globin chains, are among the most common inherited disorders. It has been shown that Hb variant remains an important cause of erroneous HbA1c results. Thus, it is important to be aware of the extent of the interference of each Hb variant encountered to avoid reporting unreliable results. However, the effects of many types of Hb variants on the measurement of HbA1c remain unclear. Here, we describe three rare Hb variants, Hb J-Tashikuergan [HBA2: c.59 C > A], Hb Pyrgos [HBB: c.251G > A], and Hb Hope [HBB: c.410 G > A], which lead to extremely high values (>25%) determined by Variant II Turbo 2.0. We further investigated their effects on HbA1c measurement by an HPLC system (Bio-Rad D100), a CE system (Sebia Capillarys 3 TERA), a boronate affinity chromatography system (Premier Hb9210), and an immunoassay method (Roche Diagnostics), and found that these Hb variants severely interfered with HbA1c measurement by Variant II Turbo 2.0 and Bio-Rad D100. This study demonstrates that patients with abnormally high HbA1c levels should be highly suspected of carrying Hb variants. When the HbA1c results are considered unreliable, other indicators such as glycated albumin may be a possible alternative to HbA1c in diabetic patients. [ABSTRACT FROM AUTHOR]

We report a clinically silent hemoglobin (Hb) variant, Hb Broomhill [α1 or α2 114(GH2) Pro > Ala;HBA1 or HBA2:c.343C > G] in a diabetic Chinese man. The Hb fractions of the subject were analyzed using various chromatographic and electrophoretic techniques. The glycated hemoglobin (HbA1c) levels measured using cation-exchange high-performance liquid chromatography (CE-HPLC) and the boronate affinity method showed nearly identical results. Analysis of the chromatogram of the CE-HPLC revealed an abnormal shoulder peak that appeared towards the end of the elution profile. Though the capillary electrophoresis method did not interpret the results, a manual examination revealed an abnormal shoulder on the HbA0 peak. Similarly, the electropherogram of the capillary zone electrophoresis also had an abnormal shoulder on the HbA peak. A missense mutation specific to the Hb Broomhill variant was found using Sanger sequencing. [ABSTRACT FROM AUTHOR]

Objectives: The US Food and Drug Administration requires donated blood to be tested for various infectious diseases to ensure safety and purity. However, testing for hemoglobin variants is not required, despite reported occurrences of hemoglobin variant transfusion and concerns about the safety of such transfusions. This study aimed to investigate the frequency of hemoglobin variants within the blood supply.Methods: We performed a 2-part study. First, we tested all RBC units in our blood bank by high-performance liquid chromatography for the presence of hemoglobin variants. Second, we performed a retrospective analysis of hemoglobin variant testing completed for routine management of sickle cell disease patients at our institution over a 5-month period to identify cases of hemoglobin variant transfusion.Results: We found that 2 of 476 (0.4%) RBC units in our blood bank contained a hemoglobin variant, and 5 of 563 (0.9%) sickle cell patients seen at our institution in a 5-month period were transfused with RBCs containing a hemoglobin variant.Conclusions: We confirmed that hemoglobin variants are present within the blood supply, and the frequency of hemoglobin variant transfusion is elevated for patients with sickle cell disease given the increased prevalence of hemoglobin variants in the population of matched donors. [ABSTRACT FROM AUTHOR]

Structural hemoglobin (Hb) variant is generally caused by a point mutation in the globin gene that produces one amino acid substitution. Here, we describe a new α-chain variant found during HbA1c measurement. HbA1c procedures based on both ion-exchange high-performance liquid chromatography (HPLC) and capillary electrophoresis (CE) techniques failed to identify its presence. In contrast, MALDI-TOF mass spectrometry (MS) revealed the existence of a variant α-chain with a mass of 15155 Da. In addition, the Hb variant interfered with HbA1c determined by Bio-Rad D100. DNA sequencing confirmed the occurrence of a new heterozygous mutation [HBA1:C.182A→G] at codon 60, resulting in a coding change from lysine to arginine. We named it Hb Liuzhou for thde birthplace of the patient. This case exemplified that MALDI-TOF mass spectrometry can serve as the method of choice to identify and quantify the Hb variant. [ABSTRACT FROM AUTHOR]

Hemoglobin (Hb) variant is one of the most common monogenic inherited disorders. We aimed to explore the prevalence and hematological and molecular characteristics of Hb variants in southern China.We collected blood samples from all patients with suspected variants found during HbA1c measurement via a cation-exchange high-performance liquid chromatography system (Bio-Rad Variant II Turbo 2.0) or a capillary electrophoresis method (Sebia Capillarys). Hematological analysis, Sanger sequencing, and gap-PCR were performed for these samples.Among the 311,024 patients tested, we found 1,074 Hb variant carriers, including 823 identified using Capillarys and 251 using Variant II Turbo 2.0, with a total carrier rate of 0.35%. We discovered 117 types of Hb variants (52 HBB, 47 HBA, and 18 HBD mutations) containing 18 new mutations. The most common variant found was Hb E, followed by Hb New York, Hb J-Bangkok, Hb Q-Thailand, Hb G-Coushatta, Hb G-Honolulu, Hb G-Taipei, and Hb Broomhill. Most heterozygotes for the Hb variant exhibited normal hematological parameters. However, most patients with compound heterozygotes for the Hb variant and thalassemia showed varied degrees of microcytic hypochromic anemia.The prevalence of hemoglobin variants remains high and exhibits genetic diversity and widespread distribution in the population of southern China. [ABSTRACT FROM AUTHOR]

Hemoglobin (Hb) North Manchester [β51(D2) Pro→His; HBB:c.155 C>A] is a rare Hb β‐globin gene variant that affects glycated Hb measurement values, such as ion‐exchange high‐performance liquid chromatography, in patients with diabetes. This variant was first detected in the UK in 1998. Here, we describe the first case involving Hb North Manchester detected incidentally in a patient with type 2 diabetes in Northern China. The Hb variant was discovered by ion‐exchange high‐performance liquid chromatography, yet capillary electrophoresis of both glycated Hb program and Hb program failed to detect it. Subsequently, Sanger sequencing was carried out to help identify the Hb variant. [ABSTRACT FROM AUTHOR]

Background Although many factors may interfere with hemoglobin (Hb)A1c measurement, Hb variants are among the most important factors. Methods We tested the HbA1c levels of the patient, a 32 year old Manchu Chinese woman, during a routine health check. We used different methods, including high-performance liquid chromatography (HPLC) and capillary electrophoresis, to test specimens from the patient. Next, we tested the specimen further using polymerase chain reaction (PCR) and sequencing. Results We discovered that our patient, who had an HbA1c value of 0, also has an Hb variant, Hb Long Island, which we found during the HbA1c analysis as part of her routine health check at the Health Management Center in the General Hospital of Tianjin Medical University, Tianjin, China. Also, we discovered that the exon 1 of β gene contained transversion mutations, with 1 heterozygous and 1 homozygous variant (HBB:c.8A > C, 9T > C). These gene mutations resulted in an amino-acid change (His to Pro) and a decrease in HbA1c value. Conclusions When there is no correlation between the clinical signs, glycemic status, and glycated Hb levels of the patient, the chromatogram of HbA1c should be carefully checked to detect possible variants that cause interference in the measurement. [ABSTRACT FROM AUTHOR]

Background: Biological diagnosis of hemoglobin disorders is a complex process relying on the combination of several analytical techniques to identify Hb variants in a particular sample. Currently, hematology laboratories usually use high-performance liquid chromatography (HPLC), capillary electrophoresis and gel-based methods to characterize Hb variants. Co-elution and co-migration may represent major issues for precise identification of Hb variants, even for the most common ones such as Hb S and C. Methods: We adapted a top-down selected reaction monitoring (SRM) electron transfer dissociation (ETD) mass spectrometry (MS) method to fit with a clinical laboratory environment. An automated analytical process with semi-automated data analysis compatible with a clinical practice was developed. A comparative study between a reference HPLC method and the MS assay was performed on 152 patient samples. Results: The developed workflow allowed to identify with high specificity and selectivity the most common Hb variants (Hb S and Hb C). Concordance of the MS-based approach with HPLC was 71/71 (100%) for Hb S and 11/11 (100%) for Hb C. Conclusions: This top-down SRM ETD method can be used in a clinical environment to detect Hb S and Hb C. [ABSTRACT FROM AUTHOR]