Evaluation of HbA1c using high performance liquid chromatography and capillary electrophoresis in Type 2 Diabetes Mellitus patients suspected to have haemoglobin variant

Abstract

BACKGROUND: High-Performance Liquid Chromatography (HPLC) is widely used for HbA1c measurement. However, it is prone to haemoglobin (Hb) variant interference. Capillary electrophoresis (CE) is believed to have better performance in patients with Hb variant. This study aimed to compare HbA1c level between HPLC and CE among Type 2 Diabetes Mellitus (T2DM) patients suspected to have Hb variant, determine the type of Hb variant among those patients, and evaluate the agreement between both methods.

METHODS: A cross-sectional study conducted at Endocrine Laboratory, Hospital Universiti Sains Malaysia, from June till December 2020. HbA1c results of adults T2DM from HPLC with suspected Hb variant were re-analysed using CE. The comparisons of HbA1c were made using paired t-test and Wilcoxon Signed Rank Test. The correlation and method comparison were made using Pearson correlation, Bland Altman (BA) and Passing-Bablok (PB), whereas the agreement using Intraclass Coefficients Correlation (ICC).

RESULTS: 250 patients were included with a median (IQR) age of 52.19 (11.11) years. For reportable results (?3.8% to ?18.5%), both methods showed no difference (p=0.382) whereas the results were difference for HbA1c >18.5% (p=0.048). 26 patients had Hb analysis with majority having Hb E trait 14 (5.6%). HPLC overestimated HbA1c in patients with Hb J and alpha Hb variant while CE able to report. Pearson correlation and PB regression analysis showed good correlation (r=0.987, p<0.001) and good agreement [slope of 1.0 (95% CI: 1.00 to 1.03); intercept of -0.3 (95% CI: ?0.61 to 0.30)]. BA plot revealed a mean difference of 0.30% (95% CI:0.00 to 0.50) with limits of agreement from ?0.54 to +1.14. ICC showed excellent reliability (0.983 (p<0.001).

CONCLUSION: HPLC and CE can be used interchangeably for HbA1c analysis across the measurement range. CE is the preferred in T2DM with certain Hb variant.