A rapid anion exchange high-performance liquid chromatography method for the measurement of HbA2 in whole blood.
We developed a binary gradient high-performance liquid chromatography (HPLC) method for measuring HbA2 in whole blood samples using a Pharmacia Mono Q column (1 mL) and measurement at 415 nm. The assay requires a simple lysis and centrifugation step before injection onto the column. We found good agreement of results between the HPLC method and the Helena column chromatography method. The within batch precision was 2-6% and between batch precision was 4-6%. We found that using 30 mM Tris buffers (pH 7-8) with a sodium chloride gradient resulted in short analysis times and good chromatographic separation of HbA2, HbS and HbA. We conclude that this is a robust assay for the diagnosis of beta-thalassaemia.
Keevil
B G
BG
Department of Clinical Chemistry, South Manchester University Hospitals NHS Trust, Wythenshawe Hospital.
Maylor
P W
PW
Rowlands
D
D
D017920:Antiporters; D016022:Case-Control Studies; D002845:Chromatography / Q000379:methods; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D006443:Hemoglobin A2 / Q000032:analysis*; D006801:Humans; D012015:Reference Standards; D015203:Reproducibility of Results; D013997:Time Factors; D017086:beta-Thalassemia / Q000097:blood / Q000175:diagnosis*
D017920:Antiporters; D006443:Hemoglobin A2