Isoelectric focusing array with immobilized pH gradient and dynamic scanning imaging for diabetes diagnosis

A traditional immobilized pH gradient (IPG) has a high stability for isoelectric focusing (IEF) but suffers from time-consuming rehydration, focusing and staining-imaging as well as complex performance. To address these issues, an IEF system with an array of 24 IPG columns (10 mm × 600 μm × 50 μm) and dynamic scanning imaging (DSI) was firstly designed for protein focusing. Moreover, two IPG columns (pH 4-9 and pH 6.7-7.7 of 10 mm in length) were firstly synthesized for IEF. A series of experiments were carried out based on the IEF array. In contrast to a traditional IPG IEF with more than 10 h rehydration, 5-14 h IEF and ca 10 h stain-imaging, the IEF array had the following merits: 25 min rehydration for sample loading, 4 min IEF, and 2 min dynamics scanning of 24 columns, well addressing the issues of traditional IEF. Furthermore, the IEF array had fair sensitivity (LOD of 60 ng), good recovery (95%), and high stability (1.02% RSD for intra-day and 2.16% for inter-day). Finally, the developed array was successfully used for separation and determination of HbA_1c (a key biomarker for diabetes diagnosis) in blood samples. All these results indicated the applicability of the developed IEF array to diabetes diagnosis.