Comparison of fast protein liquid chromatography (FPLC) with HPLC, electrophoresis & microcolumn chromatography techniques for the diagnosis of beta-thalassaemia.

BACKGROUND & OBJECTIVE: beta-thalassaemia is a genetic disorder and an important health problem around the world. Quantitative haemoglobin A(2) (HbA(2)) levels are used for the diagnosis of beta-thalassaemia. The conventional methods are high performance liquid chromatography (HPLC), electrophoresis, and microcolumn chromatography techniques. We established a fast protein liquid chromatography (FPLC) method, to measure quantitatively of HbA(2) levels, and compared its efficacy with conventional methods.

METHODS

The FPLC method, using a DEAE Sepharose, Hi Trap anion-exchange column chromatography technique was set up for HbA(2) measurement. In this study, 220 blood samples were screened for haemoglobin type by FPLC technique and also using HPLC, microcolumn chromatography and electrophoresis.

RESULTS

The FPLC results were highly correlated (r = 0.985, P<0.001) with those of HPLC for quantification of HbA(2) as well as cellulose acetate electrophoresis (r = 0.977) and microcolumn chromatography (r = 0.980). The FPLC method showed 100 per cent sensitivity and specificity, positive and negative predictive value for beta-thalassaemia diagnosis. In addition, the FPLC method was simple, rapid, low cost and reproducible. The HbA(2)/E range of FPLC for beta-thalassaemia was 6-10 per cent, HbE trait was 10-40 per cent, beta-thalassaemia/HbE was 40-60 per cent and homozygous HbE was more than 60 per cent.

INTERPRETATION & CONCLUSION: Our findings suggested that FPLC method could be used as a cost-effective method for routine beta-thalassaemia diagnosis.