Abstract
In this paper, thermally reversible hydrogel used as a replaceable packed material for capillary electrophoresis (CE) was examined. A simple and rapid method for detecting estradiol (E2) was developed, which used a strong affinity antibody (Kd=1.5×109) as a selector for immunologically based separations in serum by CE, using an uncoated capillary. Monoclonal antibody (McAb) was linked to hydrogel and used in the separation of free fluorescein isothiocyanate (FITC) labeled antigen and free antigen with laser-induced fluorescence detector (LIF). Separating conditions were optimized. E2 could be determined, specifically by capillary electrophoretic immunoassay (CEIA), without the need for pre-incubation. The N-isopropylacrylamide (PNIPA) hydrogel, including antibody, was stable and could be stored for long periods at 4°C. It can be used to determine E2 with good precision at sensitivities as low as 30pg/ml. Details of the preparation of hydrogel cross-linked monoclonal antibody and of typical separations of bound and free antigen were described.
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