Accidental detection of clinically silent compound heterozygous Hb D Punjab/Hb Q India while analyzing HbA1c by high performance liquid chromatography

HbA1c is routinely prescribed investigations for diagnosing and monitoring diabetes and high-performance liquid chromatography (HPLC) is preferred method which is also able to identify presence of hemoglobin variant. A case was encountered where presence of variant hemoglobin was indicated. On further investigation with three different instruments, diagnosis of compound heterozygous Hb D Punjab/Hb Q India was made. The chromatogram on Bio-Rad D10 showed Hb D Punjab (ααββHbD Punjab)-29.89% at 3.96 minutes retention time (RT), Hb Q India (ααHbQ Indiaββ) -9.5% with 4.45 minutes RT, hybrid of HbQ India/Hb D Punjab (ααHbQ IndiaββHbD Punjab)-6% with 4.66 minutes RT, Hb A2 (ααδδ) was 2.5% and Hb A (ααββ) was 52.2%. Analysis done on Bio-Rad variant V-II confirmed these findings. Analysis done on Sebia capillary electrophoresis revealed major peak of 50.9% in zone 9/Z(A) constituted by Hb A, second peak of 39.8% in zone 6/Z(D) constituted by co-elution of Hb D and Hb Q India, third peak of 8.8% in zone 3-4/Z(A2-C) constituted by co-elution of Hb A2 and hybrid of Hb D Punjab/Hb Q India and a fourth peak of 0.5% in zone 1 representing Hb A2HbQ India (ααHbQ Indiaδδ). Ideally variants detected while analyzing HbA1c should be further investigated for confirmation and result of which should be shared, discussed and the patient should be encouraged for screening of available family members for relevant variant hemoglobin. Combination of cation exchange HPLC and capillary electrophoresis in this case was sufficient to arrive at conclusion.

Determination of Normal and Variant Hemoglobin using Capillary Electrophoresis among Voluntary Blood Donors in North Central Nigeria: Implications on Blood Transfusion Services

Background: Voluntary non-remunerated blood donation is a strategy adopted by World Health Organization aimed at ensuring safety and adequacy of blood supply. Sub-Saharan Africa has a high prevalence of hemoglobin disorders and therefore needs to adopt stringent measures in donor selection to ensure safety for the recipient of blood transfusion. This study aimed to analyze normal and variant hemoglobin among voluntary blood donors. Methods: In this descriptive cross-sectional study, 100 prospective blood donors including 55 (55%) males and 45 (45%) females, aged 18–34 years were recruited. Capillary electrophoresis using the Minicap system was used for determining the hemoglobin variants in alkaline buffer (PH 9.4). Data analysis was done using SPSS version 20 and p-value < 0.05 was considered as the level of significance Results: The mean age of the participants was 22.23 ± 3.3 SD years. The proportion of participants with genotype AA was 67 (67%), those with AS were 17 (22 %), while those with AC were 11 (11 %). While Hb A ≥ 90% was noted in 67 (67%) blood donors, Hb S was seen in 22 (22%) and Hb A2 > 3.5% in 57 (57%). Hb F > 2% was observed in 3% of the studied participants Conclusion: Variant hemoglobin is common among blood donors and this should be taken into consideration whenever blood is being crossmatched for recipients of blood transfusion. Data from this study will be useful in raising awareness and genetic counseling. Keywords: prevalence, hemoglobin variants, capillary electrophoresis

Detection of Hemoglobinopathies by HPLC in a Referral Clinical Laboratory in Nepal

Introduction: Various hemoglobin variants are prevalent in the Nepalese population owing to the ethnic diversity of our population. Detection of asymptomatic carriers by a reliable laboratory method is the cornerstone of prevention of this serious health problem. The simplicity of the automated system with internal sample preparation, superior resolution, rapid assay time, and accurate quantification of hemoglobin fractions makes ion-exchange high-performance liquid chromatography (IEX- HPLC) an ideal methodology for the routine screening for hemoglobinopathies. We report the clinical laboratory-based prevalence of variant hemoglobin and hemoglobinopathies through IEX- HPLC analysis in a cohort of patients referred for hemoglobin electrophoresis to a referral clinical laboratory in Nepal. Materials and Methods: The variant hemoglobin and hemoglobinopathy were diagnosed based on percentage, retention time, and peak characteristics of variant hemoglobin in a chromatogram. Peripheral blood films, reticulocyte count, serum iron profile, and sickling test were done in selected cases along with detailed family history. Results: Hemoglobinopathy was detected in all age group but the vast majority was detected between 20 to 40 years of age. Beta thalassemia trait was the most frequently detected hemoglobinopathy in all age groups. Conclusions: The present study conducted using IEX-HPLC reflects the magnitude of thalassemia and hemoglobinopathies in a laboratory-based population which helps to increase awareness among both health caregivers and the general population. Routine screening for hemoglobinopathy of individuals at the reproductive age group is recommended and this screening can be done through IEX-HPLC.

A State Level Retrospective Analysis of Newborn Screening for Hemoglobinopathies

Introduction: Screening for variant hemoglobin is essential in identifying newborns born with hemoglobinopathies, including sickle cell trait (SCT). Screening allows families the opportunity for patient education as well as genetic counseling. This study aimed to better understand the trends in newborn screening of variant hemoglobin in the state of Nebraska over a period of eight years. Methods: Using the Department of Health and Human Services state-level screening information,the crude incidence rate of SCT was calculated as the screened positives divided by the total number of screened newborns within the state of Nebraska from 2011 to 2018. Rates were further delineated by variant. A Daniel's test was utilized to determine if the crude incidence rates exhibited characteristics of a trend. Rates were also examined with joinpoint regression to determine annual percent change (APC). Excel v 1808 and Joinpoint Trend Analysis v 4.5 software packages were used for all analyses; p < 0.05 was considered significant. Results: We identified 3,416 newborns with SCT among a total of 211,697 live births, which is an incidence rate of 161.4 cases per 10,000 screened newborns. There was no evidence of a trend in the overall incidence rate from 2011 to 2018 (p = 0.493). However, the incidence of FA Barts significantly increased over the examined interval (p = 0.015) with a 4.4% APC (95% CI: 0.7% to 8.3%; p =0.026) Conclusion: The data obtained from this study showed no significant change in the incidence of SCT in the state of Nebraska from 2011 to 2018. However, there was a significant increase in the incidence of Hemoglobin FA Barts. Hemoglobin FA Barts is associated primarily with alpha thalassemia, which is most prevalent in southeast Asia, but is also common in the Mediterranean, Africa, Middle east, Central Asia and India. This finding is of interest and likely related to the changing migration patterns in the state of Nebraska. For the healthcare system to continue to provide appropriate care, future studies should aim to further elucidate this trend and patient demographics of this increasing patient population in the state of Nebraska. Disclosures No relevant conflicts of interest to declare.

Evaluation of efficacy of BIORAD D10TM testing system in detection of beta thalassaemia carrier

Background: Hemoglobinopathies especially thalassaemia and its interaction with HbE and HbS are significant cause of morbidity and mortality in our country. With no feasible treatment, prevention of cases by carrier detection is the only option for successful reduction of the disease burden. VARIANT hemoglobin testing system BIORAD using principle of cation exchange chromatography has been in use and considered as gold standard for carrier detection. The purpose of this study is to compare the efficacy of a different, cheaper instrument; D10 by the same manufacturer BIORAD for carrier detection in beta thalassaemia.Methods: Patients diagnosed as beta thalassaemia carrier by VARIANT hemoglobin testing system (HbA2 value between 4.0-9.0) were retested using D10 instrument and checked for agreement.Results: There was good correlation between VARIANT and D-10 methods with Intraclass correlation coefficient 0.832 (95% Confidence Interval 0.756-0.884). Bland-Altman analysis showed mean bias of +0.3526 (95% CI -0.3958 to +1.101).Conclusions: Although further study is needed with larger sample size for assessment of sensitivity and specificity of D10 instrument, it is evident from this study that this instrument can be an effective and cheaper alternative of VARIANT hemoglobin testing system.