Methods, units and quality requirements for the analysis of haemoglobin A1c in diabetes mellitus

Abstract

The formation of glycohemoglobin, especially the hemoglobin A_1c (HbA_1c) fraction, occurs when glucose becomes coupled with the amino acid valine in the β-chain of Hb; this reaction is dependent on the plasma concentration of glucose. Since the early 1970s it has been known that diabetics display higher values OF HbA_1C because they have elevated blood glucose concentrations. Thus HbA_1c has acquired a very important role in the treatment and diagnosis of diabetes mellitus. After the introduction of the first quantitative measurement OF HbA_1C, numerous methods for glycohemoglobin have been introduced with different assay principles: From a simple mini-column technique to the very accurate automated high-pressure chromatography and lastly to many automated immunochemical or enzymatic assays. In early days, the results of the quality control reports for HbA_1c varied extensively between laboratories, therefore in United States and Canada working groups (WG) of the Diabetes Controls and Complications Trial (DCCT) were set up to standardize the HbA_1c assays against the DCCT/National Glycohemoglobin Standardization Program reference method based on liquid chromatography. In the 1990s, the International Federation of Clinical Chemistry and Laboratory Medicine (IFCC) appointed a new WG to plan a reference preparation and method for the HBA_1c measurement. When the reference procedures were established, in 2004 IFCC recommended that all manufacturers for equipment used in HbA_1c assays should calibrate their methods to their proposals. This led to an improvement in the coefficient of variation (CV%) associated with the assay. In this review, we describe the glycation of Hb, methods, standardization of the HbA_1c assays, analytical problems, problems with the units in which HbA_1c values are expressed, reference values, quality control aspects, target requirements for HbA_1c, and the relationship of the plasma glucose values to HbA_1c concentrations. We also note that the acceptance of the mmol/mol system for HbA_1c as recommended by IFCC, i.e., the new unit and reference ranges, are becoming only slowly accepted outside of Europe where it seems that expressing HbA_1c values either only in per cent units or with parallel reporting of percent and mmol/mol will continue. We believe that these issues should be resolved in the future and that it would avoid confusion if mmol/mol unit for HbA_1c were to gain worldwide acceptance.

Keywords: Diabetes; Hemoglobin A_1c; Glycohemoglobin; Glucose; International Federation of Clinical Chemistry and Laboratory Medicine; Reference values; Quality assurance; Recommendation; Target limits

Core tip: The aim of this review is to clarify methods, units, quality requirements, reference and cutoff limits for hemoglobin A_1c (HbA_1c) and ratio of blood glucose/HbA_1c on the basis of the results from Finnish quality control surveys by comparing them to the literature. The HbA_1c surveys of Labquality Ltd. (Helsinki, Finland) were started in 1986 by using two fresh EDTA-blood samples. From 1994, the number of the participating laboratories had risen to 139, of which 75 were Finnish and 64 from five other countries. In 2014, the number of the participating laboratories was total 214, 141 were Finnish and 73 from 13 other countries.