Evaluation of effects from hemoglobin variants on HbA1c measurements by different methods.

OBJECTIVES: The impact of seven hemoglobin variants (Hb Q-Thailand, Hb G-Honolulu, Hb Ube-2, Hb New York, Hb J-Bangkok, Hb G-Coushatta, and Hb E) on the outcome of HbA1c was investigated for six methods by comparing with liquid chromatography-tandem mass spectrometry (LC/MS/MS) reference method. METHODS: Twenty-nine normal and 112 variant samples were measured by LC/MS/MS, Sebia Capillarys 3 TERA, Intelligene Biosystems QuanTOF, Premier Hb9210, Arkray HA-8190V, Bio-Rad D-100, and Tosoh G11, then evaluated for correlation, consistency, and mean relative bias among six methods. The lowest biological variation bias of ±2.8 % was an acceptable standard. RESULTS: All methods showed poor correlation and consistency with LC/MS/MS for Hb E. The unacceptable biases were observed for Capillarys 3 TERA (-14.4 to -3.7 % for Hb Q-Thailand, Hb Ube-2, Hb New York, Hb J-Bangkok and Hb E), QuanTOF (-8.3 to -2.9 % for Hb Ube-2, Hb New York and Hb G-Coushatta), Premier Hb9210 (-18.3 to -3.6 % for Hb Q-Thailand, Hb Ube-2, Hb New York, Hb J-Bangkok and Hb E), HA-8190V variant mode (-17.3 to 6.6 % for Hb G-Honolulu, Hb Ube-2, Hb New York, Hb G-Coushatta and Hb E). All variant samples showed larger biases than ±2.8 % comparing HA-8190V fast mode, D-100, and G11 with LC/MS/MS. CONCLUSIONS: The accuracy of different HbA1c methods was influenced by some Hb variants, especially Hb Ube-2 and Hb New York. Thus, laboratories need to choose appropriate methods to measure HbA1c with different Hb variants.

Wide spectrum of novel and rare hemoglobin variants in the multi-ethnic Indian population: A review.

The hemoglobin (Hb) variants are qualitative abnormalities due to production of structurally abnormal globin proteins. They are categorized based on the type of mutation present in the α1, α2, ß, Gγ, Aγ and δ globin genes. So far, more than 1550 Hb variants are reported in the database. They could lead to Hb polymerization, Hb instability, altered oxygen affinity and decreased oxygen-carrying capacity of Hb or have no clinical manifestations. In India, ethnic diversity, consanguinity, regional variations and migration result in the presence of different Hb variants. We have compiled all the variants of α, ß and δ globin chains in heterozygous, homozygous and in compound heterozygous forms reported from India in the last 52 years. Of the 63 rare and novel hemoglobin variants reported from India, 22 were α-globin chain variants, 37 were ß-globin chain variants and 4 were δ-globin chain variants. Twelve novel Hb variants (Hb J Rajappan, Hb Koya Dora, Hb Rampa, Hb Godavari, Hb Chandigarh, Hb D Agri, Hb Lucknow, Hb Vellore, Hb Midnapore, Hb Bijnor, Hb A2Tianhe and Hb A2Saurashtra) were identified among persons of Indian origin. Majority of them were picked up on HPLC. Some of the variants like Hb Titusville, Hb Shimonoseki, Hb Chandigarh, Hb D Agri, Hb Yaizu and Hb Vellore eluted in the HbS window whereas variants like HbD Iran, Hb St. Louis, Hb G Coushata, HbM Saskatoon, Hb Lucknow, Hb Grange-Blanche and Hb Tianshui showed falsely elevated HbA2. Hence, careful and systematic investigations are required to identify them.

Point-of-Care Diagnostic Test for Beta-Thalassemia.

Hemoglobin (Hb) disorders are among the most common monogenic diseases affecting nearly 7% of the world population. Among various Hb disorders, approximately 1.5% of the world population carries ß-thalassemia (ß-Thal), affecting 40,000 newborns every year. Early screening and a timely diagnosis are essential for ß-thalassemia patients for the prevention and management of later clinical complications. However, in Africa, Southern Europe, the Middle East, and Southeast Asia, where ß-thalassemia is most prevalent, the diagnosis and screening for ß-thalassemia are still challenging due to the cost and logistical burden of laboratory diagnostic tests. Here, we present Gazelle, which is a paper-based microchip electrophoresis platform that enables the first point-of-care diagnostic test for ß-thalassemia. We evaluated the accuracy of Gazelle for the ß-Thal screening across 372 subjects in the age range of 4-63 years at Apple Diagnostics lab in Mumbai, India. Additionally, 30 blood samples were prepared to mimic ß-Thal intermediate and ß-Thal major samples. Gazelle-detected levels of Hb A, Hb F, and Hb A2 demonstrated high levels of correlation with the results reported through laboratory gold standard high-performance liquid chromatography (HPLC), yielding a Pearson correlation coefficient = 0.99. This ability to obtain rapid and accurate results suggests that Gazelle may be suitable for the large-scale screening and diagnosis of ß-Thal.

First report of a patient with homozygous hemoglobin Ernz: Evidence to support a non-pathogenic variant.

Hemoglobin Ernz (Hb Ernz) is a missense variant in ß-globin caused by a Threonine to Asparagine substitution at the 123rd amino acid position and HBB c.371C > A in gene level. Hb Ernz has been classified as Uncertain Significance (VUS) by ACMG due to limited reports and the absence of any homozygote genotypes. In our study, we found eight cases of Hb Ernz by DNA sequencing of the ß-globin gene during >20 years of Thalassemia Screening in individuals with borderline hematological parameters who were possible carriers of thalassemia or their spouses. We also report the first homozygote variant of Hb Ernz. Our findings suggest that the changes in hematological parameters observed in individuals with Hb Ernz are likely due to α-globin gene mutations rather than Hb Ernz itself. These findings support the reclassification of Hb Ernz as a benign variant in variant classification.

Evaluation of the Premier Hb9210 instrument for HbA1c determination.

Background: Glycated hemoglobin measurements are a valuable tool for long-term blood glucose monitoring and the diagnosis of diabetes. Its widespread use has been made possible due to the development of new analytical methods with improved performances and standardization with reference materials. The aim of the present study was to evaluate the Trinity Biotech Premier Hb9210 analyzer for the measurement of HbA1c.Methods: The precision was assessed using the CLSI EP-15A3 and EP-10A3 protocols. The latter was also used to investigate linearity, carryover, and linear drift. The comparison study was performed between Premier Hb910 and Tosoh HLC-723 G8 through Passing-Bablok regression and the Bland-Altman plot. The Fleiss Kappa index was used to assess the degree of agreement. The interference of Hb variants was investigated using samples with Hb variants S, C, D, E, J, and Seville.Results: Within-run and between-run imprecision fell between 0.37% and 1.16%. No statistically significant nonlinearity, carry-over, and/or drift were observed. The resulting regression line of the Passing-Bablok analysis was y = 0.00 + 1.00x. The Pearson correlation coefficient was 0.997. In the Bland-Altman plot, the relative bias was 0.01%. The overall Fleiss Kappa index was 0.9. No interference from hemoglobin variants was observed.Conclusion: The Premier Hb9210 demonstrated a high degree of automation, reproducibility, good agreement, minimal carry-over effect, and excellent linearity across the wide range of HbA1c levels commonly found in diabetic patients and was not influenced by Hb variants.

Premier Hb9210 can be used as an alternative to monitor glycemic status.Premier Hb9210 is not affected by common hemoglobin variants.Premier Hb9210 can correctly classify diabetic patients.

Association between hemoglobin variants and laboratory outcomes in patients infected with P. falciparum from South West Uganda.

Aims: We assess the relationship between various hemoglobin variants and some hematological parameters packed cell volume, white blood cells (PCV, WBC) and parasitemia level of patients with malaria in the southwestern, Uganda. Methods: Patient were enrolled by rapid diagnostic tests (RDTs), confirmed by microscopy, and laboratory outcomes were determined. Results: Patients positive for malaria RDTs were 155, microscopic-confirmed P. falciparum parasites were 95 (61.29%) having hemoglobin variants HbAA and HbAS; 75 (78.95%) and 13 (13.68%), respectively. The laboratory outcomes showed mean, PCV (32.19 ± 4.83), WBC (5831.66 ± 2888.29) and P. falciparum parasitaemia density (32,605.45 ± 14031), while the hemoglobin variants mean values AA (39,008.85 ± 31,261.56), AC (15908 ± 10173.48), AS (16,561.46 ± 15,380.93), SC (30,524 ± 0.000) and SS(1652 ± 0.000) were significantly different from the total population (34,321.5 ± 21,924.26) parasite-density. Conclusion: Patients with hemoglobin variants HbAA had a significantly higher parasite-carrying capacity and PCV levels.

Hemoglobin Variants in Patients With Microcytic Hypochromic Anemia: A Review of Indian Studies.

Microcytic hypochromic (MCHC) anemia with hemolytic components is common in clinical practice. Hemoglobinopathies and variants are one of the important underlying causes of MCHC anemia. The Indian population, by large, as various studies reported, showed a plethora of hemoglobinopathies with regional predilections for its types. The present systematic review is carried out for the evaluation of MCHC anemia for its underlying causes of hemoglobinopathies and their loco regional comparisons. The review was carried out by the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) method with selected keywords through the Google advanced search matchable to the objectives of the present systematic review. Upon the systematic review, it was observed that ß thalassemia trait (ßTT) remained the highest reported hemoglobinopathy. The other abnormal hemoglobin variants, though rare, also have been reported in the reviewed articles. It is concluded that patients with MCHC refractory to its regular treatment should be subjected to high-performance liquid chromatography (HPLC) in exclusion of underlying hemoglobinopathy and abnormal hemoglobin variants.

Quantitation of hemoglobins using Sebia Capillarys-2 capillary electrophoresis (CE) for A1c: Comparison to results using CE for hemoglobins.

Background: Measurement of A1c using the Sebia Capillarys-2 capillary electrophoresis (A1c CE) involves relative quantitative measurements of peaks for hemoglobins A1c, A, A2. We examined correlation of A1c CE results with results of CE analysis for hemoglobins (Hb CE) for homozygous A and S-trait patients. We specifically examined whether abnormalities in A2 or the A/S ratio by A1c CE alone would reasonably be the basis for recommendation of red cell indices for evaluation of possible thalassemia. Methods: Selection of patients was from results for A1c CE, exhibiting either a normal pattern or a pattern consistent with S-trait. We then examined correlation of results of quantitation for A, S and A2 between A1c CE and Hb CE. Results: %A2 by A1c CE (y) had high correlation with %A2 by Hb CE (x): y = 0.88 x; r = 0.948. %A2 in S-trait patients was right-shifted in comparison to normals by 0.5%. For S-trait patients, the A/S ratio by A1c CE (y) had high correlation with the A/S ratio by Hb CE (x): y = 1.02 x; r = 0.995. Conclusions: Given high correlation of results between A1c CE and Hb CE, patent elevation of A2 by A1c CE for either normal or S-trait patients is a reasonable basis for recommendation of red cell indices for evaluation of possible beta thalassemia. For S-trait patients, patent abnormality in the A/S ratio by A1c CE is a reasonable basis for recommendation of red cell indices for evaluation of possible alpha or beta thalassemia.

Epidemiological Survey of Hemoglobinopathies Based on Next-Generation Sequencing Platform in Hunan Province, China.

Objective: This study was aimed at investigating the carrier rate of, and molecular variation in, α- and ß-globin gene mutations in Hunan Province. Methods: We recruited 25,946 individuals attending premarital screening from 42 districts and counties in all 14 cities of Hunan Province. Hematological screening was performed, and molecular parameters were assessed. Results: The overall carrier rate of thalassemia was 7.1%, including 4.83% for α-thalassemia, 2.15% for ß-thalassemia, and 0.12% for both α- and ß-thalassemia. The highest carrier rate of thalassemia was in Yongzhou (14.57%). The most abundant genotype of α-thalassemia and ß-thalassemia was -α 3.7/αα (50.23%) and ß IVS-II-654/ß N (28.23%), respectively. Four α-globin mutations [CD108 (ACC>AAC), CAP +29 (G>C), Hb Agrinio and Hb Cervantes] and six ß-globin mutations [CAP +8 (C>T), IVS-II-848 (C>T), -56 (G>C), beta nt-77 (G>C), codon 20/21 (-TGGA) and Hb Knossos] had not previously been identified in China. Furthermore, this study provides the first report of the carrier rates of abnormal hemoglobin variants and α-globin triplication in Hunan Province, which were 0.49% and 1.99%, respectively. Conclusion: Our study demonstrates the high complexity and diversity of thalassemia gene mutations in the Hunan population. The results should facilitate genetic counselling and the prevention of severe thalassemia in this region.

Evaluation of local hemoglobinopathy prevalence and promotion of accurate hemoglobin A1c testing using historical data retrieval.

OBJECTIVES: Heterozygous hemoglobin variants are known to cause method- and variant-specific interference with hemoglobin A1c (HbA1c) quantitation. Less attention has been paid to the role of other hemoglobin variants in confounding HbA1c testing. Here we evaluated the frequency with which enzymatic (ENZ) and immunoassay (IA) HbA1c quantitation methods, i.e., those unable to routinely detect the presence of hemoglobin variants, were used within our healthcare system for HbA1c analysis in patients with elevated fetal hemoglobin as well as compound heterozygous and homozygous variants. DESIGN & METHODS: This analysis was enabled by automated review of HbA1c result history, implemented to promote detection of variants prior to HbA1c result reporting. RESULTS: During a 54-week period, 319,290 HbA1c analyses were performed. We observed 110 unique patient cases (0.03% problem identification rate) in which HbA1c testing was ordered in the presence of either a homozygous or compound heterozygous hemoglobin variant or elevated hemoglobin F beyond the tolerance of the method. Among the 110 cases identified, 55 (50%) showed a compound heterozygous or homozygous hemoglobin variant while 55 (50%) showed elevated hemoglobin F. Of those cases involving a compound heterozygous or homozygous variant, 8/55 (15%) involved patients who had one or more ENZ or IA HbA1c results reported previously within our system. Of the 55 total compound heterozygous or homozygous variants identified, 37 (67%) were hemoglobin E, 10 (18%) hemoglobin S/C, 4 (7%) hemoglobin S, 2 (4%) hemoglobin C, 1 (2%) hemoglobin Camden, and 1 (2%) unidentified variant. CONCLUSIONS: Exclusive use of methods unable to routinely detect the presence of hemoglobin variants may lead to reporting of HbA1c results that are not clinically meaningful.

Red Blood Cell Substitutes: Liposome Encapsulated Hemoglobin and Magnetite Nanoparticle Conjugates as Oxygen Carriers.

The established blood donation and transfusion system has contributed a lot to human health and welfare, but for this system to function properly, it requires a sufficient number of healthy donors, which is not always possible. Pakistan was a country hit hardest by COVID-19 which additionally reduced the blood donation rates. In order to address such challenges, the present study focused on the development of RBC substitutes that can be transfused to all blood types. This paper reports the development and characterization of RBC substitutes by combining the strategies of conjugated and encapsulated hemoglobin where magnetite nanoparticles would act as the carrier of hemoglobin, and liposomes would separate internal and external environments. The interactions of hemoglobin variants with bare magnetite nanoparticles were studied through molecular docking studies. Moreover, nanoparticles were synthesized, and hemoglobin was purified from blood. These components were then used to make conjugates, and it was observed that only the hemoglobin HbA1 variant was making protein corona. These conjugates were then encapsulated in liposomes to make negatively charged RBC substitutes with a size range of 1-2 µm. Results suggest that these RBC substitutes work potentially in a similar way as natural RBCs work and can be used in the time of emergency.

Interference of hemoglobin variants in HbA1c quantification.

Fasting blood glucose and glycated hemoglobin (HbA1c) are routine biomarkers to screen and monitor diabetes mellitus. HbA1c results from glycation at the N-terminus of the ß globin chain of tetrameric human hemoglobin. Fasting blood glucose level varies with the nature and amount of food intake, physical exercise, etc., and, accordingly, is a short-term measure of glucose control. In contrast, HbA1c provides an average measure of glucose control for the long-term (8-12 weeks). Unfortunately, genetic variants of hemoglobin may interfere with HbA1c quantification using ion exchange chromatography, capillary electrophoresis, immunoassay and boronate affinity chromatography. Mass spectrometry, however, measures total glycation of hemoglobin across both α and ß globin chains and correlates well with the ion exchange based method. Additionally, mass spectrometry based quantification is not impacted by the presence of genetic variants of hemoglobin and thus might be a better analytical choice for diabetes mellitus.

Epidemiological Survey of Hemoglobinopathies Based on Next-Generation Sequencing Platform in Hunan Province, China / 生物医学与环境科学（英文）

OBJECTIVE@#This study was aimed at investigating the carrier rate of, and molecular variation in, α- and β-globin gene mutations in Hunan Province.@*METHODS@#We recruited 25,946 individuals attending premarital screening from 42 districts and counties in all 14 cities of Hunan Province. Hematological screening was performed, and molecular parameters were assessed.@*RESULTS@#The overall carrier rate of thalassemia was 7.1%, including 4.83% for α-thalassemia, 2.15% for β-thalassemia, and 0.12% for both α- and β-thalassemia. The highest carrier rate of thalassemia was in Yongzhou (14.57%). The most abundant genotype of α-thalassemia and β-thalassemia was -α 3.7/αα (50.23%) and β IVS-II-654/β N (28.23%), respectively. Four α-globin mutations [CD108 (ACC>AAC), CAP +29 (G>C), Hb Agrinio and Hb Cervantes] and six β-globin mutations [CAP +8 (C>T), IVS-II-848 (C>T), -56 (G>C), beta nt-77 (G>C), codon 20/21 (-TGGA) and Hb Knossos] had not previously been identified in China. Furthermore, this study provides the first report of the carrier rates of abnormal hemoglobin variants and α-globin triplication in Hunan Province, which were 0.49% and 1.99%, respectively.@*CONCLUSION@#Our study demonstrates the high complexity and diversity of thalassemia gene mutations in the Hunan population. The results should facilitate genetic counselling and the prevention of severe thalassemia in this region.

Interpretation of HbA1c lies at the intersection of analytical methodology, clinical biochemistry and hematology (Review).

Over past few decades, diabetes has become widespread on a global scale. Hemoglobin A1c (HbA1c) assessment is crucial for diabetes care, since it allows for the monitoring of an individual's level of glycemic control over the course of 2 to 3 months and risk assessment to determine any possible complications. Numerous methods, including cation-exchange chromatography, electrophoresis, immunoassays and affinity chromatography, can be used to determine the HbA1c level. Each method has its limitations, however. The amount of HbA1c in patient samples is not only dependent on blood glucose levels, but is also strongly influenced by changes in red blood cell lifespan and globin chain structure. Consequently, hematological, clinical biochemistry and analytical methods all intertwine when interpreting HbA1c. There are numerous reports on the interactions of HbA1c with inherited and acquired diseases. Some of these impacts are inconsistent and difficult to explain. The present review article aimed to summarize and classify these effects and evaluate their clinical relevance. The findings discussed herein may serve as a reminder that clinical HbA1c values need to be analyzed with caution.

Optimization and Identification of Single Mutation in Hemoglobin Variants with 2,2,2 Trifluoroethanol Modified Digestion Method and Nano-LC Coupled MALDI MS/MS.

Background: Hemoglobin (Hb) variants arise due to point mutations in globin chains and their pathological treatments rely heavily on the identification of the nature and location of the mutation in the globin chains. Traditional methods for diagnosis such as HPLC and electrophoresis have their own limitations. Therefore, the present study aims to develop and optimize a specific method of sample processing that could lead to improved sequence coverage and analysis of Hb variants by nano LC-MALDI MS/MS. Methods: In our study, we primarily standardized various sample processing methods such as conventional digestion with trypsin followed by 10% acetonitrile treatment, digestion with multiple proteases like trypsin, Glu-C, Lys-C, and trypsin digestion subsequent to 2,2,2 trifluoroethanol (TFE) treatment. Finally, the peptides were identified by LC-MALDI MS/MS. All of these sample processing steps were primarily tested with recombinant Hb samples. After initial optimization, we found that the TFE method was the most suitable one and the efficiency of this method was applied in Hb variant identification based on high sequence coverage. Results: We developed and optimized a method using an organic solvent TFE and heat denaturation prior to digestion, resulting in 100% sequence coverage in the ß-chains and 95% sequence coverage in the α-chains, which further helped in the identification of Hb mutations. A Hb variant protein sequence database was created to specify the search and reduce the search time. Conclusion: All of the mutations were identified using a bottom-up non-target approach. Therefore, a sensitive, robust and reproducible method was developed to identify single substitution mutations in the Hb variants from the sequence of the entire globin chains. Biological Significance: Over 330,000 infants are born annually with hemoglobinopathies and it is the major cause of morbidity and mortality in early childhood. Hb variants generally arise due to point mutation in the globin chains. There is high sequence homology between normal Hb and Hb variant chains. Due to this high homology between the two forms, identification of variants by mass spectrometry is very difficult and requires the full sequence coverage of α- and ß-chains. As such, there is a need for a suitable method that provides 100% sequence coverage of globin chains for variant analysis by mass spectrometry. Our study provides a simple, robust, and reproducible method that is suitable for LC-MALDI and provides nearly complete sequence coverage in the globin chains. This method may be used in the near future in routine diagnosis for Hb variant analysis.

Monitoring of glycemic balance by glycated hemoglobin assay on D-10® (Biorad) and Minicap Flex Piercing® (Sebia) biochemical systems at the Rodolph-Mérieux Laboratory in Bamako / Suivi de l'équilibre glycémique par le dosage de l'hémoglobine glyquée sur les automates de biochimie D-10® (Biorad) et Minicap Flex Piercing® (Sebia) au Laboratoire RodolpheMérieux de Bamako

This study aims to determine the concordance of results between D-10® and Minicap Flex Piercing® and the impact of the presence of hemoglobin variants in the glycated hemoglobin assay. All the samples were assayed in parallel. The agreement between these methods was assessed using the Bland and Altman plot. We collected a total of 166 samples. In the entire study population and in patients with a hemoglobin variant, the Bland and Altman graph showed good agreement between the values (for respective biases of 0.21% and 0.29%) as well as a significant correlation (p < 0.001). Our results differ from those found by Kesso Barry et al. This difference can be explained by the high prevalence of patients with abnormal hemoglobins S and C (39.2 %) in our study population. Despite a good agreement between the methods, the results do not allow us to confirm a transferability between the two systems in diabetics and carriers of hemoglobin variants.

Notre étude vise à déterminer la concordance des résultats entre le D-10® et le Minicap Flex Piercing®, ainsi que l'impact de la présence des variants d'hémoglobine dans le dosage de l'hémoglobine glyquée. Tous les échantillons ont été dosés en parallèle sur les deux automates. L'évaluation de la concordance a été réalisée grâce au diagramme de Bland et Altman. Nous avons colligé un total de 166 échantillons. Chez l'ensemble de la population d'étude et chez les patients avec un variant d'hémoglobine, le graphique de Bland et Altman a montré une bonne concordance entre les valeurs (pour des biais respectifs de 0,21 % et 0,29 %) ainsi qu'une corrélation significative (p < 0,001). Nos résultats diffèrent de ceux trouvés par Kesso Barry et al. Cette différence peut s'expliquer par la forte prévalence de patients possédant des hémoglobines anormales S et C (39,2 %) dans notre population d'étude. Malgré une bonne concordance, nos résultats ne permettent pas d'affirmer une transférabilité entre les deux systèmes chez les diabétiques et les porteurs de variants d'hémoglobine.

An overview of sickle cell disease from the socio-demographic triangle - a Nigerian single-institution retrospective study.

Introduction: Sickle cell disease (SCD) is a hereditary red blood cell disorder of public health importance globally with Nigeria the epicenter zone in Africa. There is a paucity of knowledge on how hemoglobin variants, personal characteristics, and environment (socio-demographic triangle) interact to influence SCD propagation. A clinical overview of these epidemiologic parameters may proffer strategies for controlling the SCD disease burden. The objective of this study was to examine the prevalence patterns of SCD, including other associated epidemiologic and hematological (i.e., hemoglobin concentration, ABO blood groups) parameters from laboratory data. Methods: this was a retrospective cross-sectional study of 138 newly diagnosed SCD patients in the laboratory unit of the department of haematology using routine alkaline cellulose acetate hemoglobin electrophoresis technique from 2013 to 2014. Demographic and other relevant data were obtained from case notes and laboratory records at the presentation. The agent-host-environment variables were used in the construction of the epidemiological triad chain of transmission. Results: a total of 138 (1.63%) newly diagnosed SCD patients aged 7 months to 41 years made up of 39% (0.63% SCD prevalence) adults and 61% (1% SCD prevalence) pediatric age-groups were seen out of 8457 consecutive patients screened within the study period. About 98.55% and 1.45% were homozygous sickle-hemoglobin (SS) and heterozygous sickle-hemoglobin C (SC) variants, respectively. The pediatric department (CHER+CHOP) recorded the highest proportion of SCD (65%). In contrast, the public health department had the least proportion (1%). There was a statistically significant difference in the gender status and the months of SCD diagnosis (p=0.0147). The month of April had the highest proportion of SCD. A majority (66.7%) of the SCD had moderate grade anemia. Conclusion: the study revealed a gender disparity in the months of SCD diagnosis. However, there was no statistical difference in the pediatric and adult SCD prevalence patterns.

Evaluation of Volumetric Absorptive Microsampling and Mass Spectrometry Data-Independent Acquisition of Hemoglobin-Related Clinical Markers.

Data-independent acquisition (DIA) allows comprehensive proteome coverage, while it also potentially works as a unified protocol to determine a multitude of proteins found in blood. Because of its high specificity, mass spectrometry may greatly reduce the interference observed in other assays to evaluate blood markers. Here, we combined DIA with volumetric absorptive microsampling (VAMS) and automated proteomics sample processing in a platform to assess clinical markers. As a proof of concept, we evaluated two hemoglobin-related biomarkers: the glycated hemoglobin (HbA1c) and hemoglobin (Hb) variants. HbA1c by DIA showed good correlation with the reference method, but method imprecision did not meet the quality requirement for this biomarker. We developed a strategy to identify Hb variants based on a customized database combined with a workflow for DIA data extraction and rigorous peptide evaluation. Data are available via ProteomeXchange with identifier PXD029918.