On-line microheterogeneity analysis and rapid phenotyping of haptoglobin by capillary electrophoresis using sodium...

To improve the detection sensitivity and determine phenotypes of haptoglobin (Hp), a prefilling technique was developed and tested in capillary electrophoresis (CE) with UV-vis absorbance detection. Adding 0.01% sodium dodecyl sulfate (SDS) to the protein sample and 0.1% SDS to the prefilling buffer solution, on-line stacking and microheterogeneity separation of Hp were achieved. In addition, the influences of pH, buffer concentration, sample and prefilling buffer SDS concentration upon resolution were examined. Under optimized conditions, Hp-microheterogeneity was well resolved and two phenotypes of Hp (Hp 1-1 and Hp 2-2) were differentiated. This method was applied to the analysis of sera from normal individuals and beta-Thalassemia patients. After the depletion of albumin (HSA) and immunoglobulin G (IgG), this method allowed to determine two phenotypes in different individuals and to detect the decrease of Hp in beta-Thalassemia patients. Featuring high efficiency, speed and simplicity, the proposed method shows great potential for use in clinical diagnosis and proteome research.