Unknown Peaks and Spuriously Low Values of Glycated Haemoglobin by High Performance Liquid Chromatography: A Cross-sectional Survey

S. Chakraborty
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Abstract

Introduction: Glycated haemoglobin (HbA1c) has humongous role both in diagnosis as well as in guiding the clinicians in making therapeutic decision in diabetic patients. There are various methods of laboratory estimation of glycated haemoglobin depending on charge and affinity. However, the methods are affected by a number of factors like haemoglobin variants, drugs and erythrocyte survival. Aim: To identify and evaluate abnormal peaks in chromatogram of High Performance Liquid Chromatography (HPLC) and compare it with boronate affinity chromatography and it also assessed the interference of elution of silent haemoglobin variants or drugs in exactness of HbA1c estimation by HPLC. Materials and Methods: A cross-sectional survey was conducted in the laboratory of Purwanchal Nagarik Samity, Kolkata, West Bengal, India, from November 2020 to August 2021, where 2500 samples were evaluated for HbA1c estimation. Abnormal variant window, unknown peaks or spuriously low HbA1c were identified in eight samples, where estimation was done by HPLC. These were processed further to investigate for the haemoglobin variants. The glycated haemoglobin in those samples was further estimated by Boronate affinity chromatography. Results: Out of the eight samples, three patients were Haemoglobin E trait, two were Haemoglobin E homozygous and one had sickel cell trait. However, two samples showed a normal chromatogram in HbA2/HbF mode. The glycated haemoglobin was affected by Haemoglobin E, sickel cell traits as well as other interferences that may cause inappropriate lowering of glycated haemoglobin. Conclusion: The results of this study and literature review suggests the progression of various micro/macrovascular complications in diabetic individuals can be arrested by maintaining normal HbA1c levels. Therefore, the laboratory consultant should meticulously take into consideration factors like plasma glucose concentration, drug history, biological variation or abnormal haemoglobinopathies. Thus, every laboratory catering to the huge burden of diabetic patients, where the prevalence of haemoglobinopathy is high, should individualise the method of HbA1c estimation.
用高效液相色谱法测定糖化血红蛋白的未知峰和伪低值:横断面调查
导读:糖化血红蛋白(HbA1c)在糖尿病患者的诊断和指导临床医生的治疗决策中都具有巨大的作用。根据电荷和亲和力,有各种各样的实验室估计糖化血红蛋白的方法。然而,这些方法受到许多因素的影响,如血红蛋白变异、药物和红细胞存活。目的:鉴别和评价高效液相色谱(HPLC)中异常峰,并将其与硼酸盐亲和色谱进行比较,同时评价无症状血红蛋白变异或药物洗脱对HPLC测定HbA1c准确性的干扰。材料和方法:2020年11月至2021年8月,在印度西孟加拉邦加尔各答Purwanchal Nagarik Samity实验室进行了一项横断面调查,对2500个样本进行了HbA1c估计评估。在8个样本中发现了异常变异窗口、未知峰或假低的HbA1c,并通过HPLC进行了估计。这些被进一步处理以调查血红蛋白变异。通过硼酸盐亲和层析进一步估计了这些样品中的糖化血红蛋白。结果:8例患者中,3例为血红蛋白E性状,2例为血红蛋白E纯合性状,1例为镰状细胞性状。然而,两个样品在HbA2/HbF模式下显示正常的色谱图。糖化血红蛋白受到血红蛋白E、镰状细胞特性以及其他可能导致糖化血红蛋白不适当降低的干扰的影响。结论:本研究结果和文献综述表明,维持正常的HbA1c水平可以阻止糖尿病患者各种微血管/大血管并发症的进展。因此,实验室会诊医师应仔细考虑血糖浓度、用药史、生物变异或异常血红蛋白病等因素。因此,每个满足糖尿病患者巨大负担的实验室都应该个性化HbA1c估计方法,因为糖尿病患者的血红蛋白病患病率很高。
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