Dielectrophoretic detection of molecular bindings

The specificity of molecular binding between the "target" and the "probe" molecule, for example, between antigen and antibody or between two complementary deoxyribonucleic acid (DNA) sequences, is the principle of affinity assays. In the assay, the target is mixed with the fluorescence-labeled probe, so that the probe binds to the target to form a target-probe complex. Then, the bound complex is separated from the free (unbound) probe somehow (bound/free (BF) separation), and the fluorescence emission from the separated complex is measured to obtain the target concentration in the original sample. In this paper, the authors propose and experimentally demonstrate the use of dielectrophoresis (DEP) for such B/F separations. Using DEP chromatography, DEP characteristics of various biomolecules are measured, and: (1) separation of A-DNA (48.5 kbp) and oligonucleotide (22base); and (2) quantitative detection of antigen-antibody bindings, are demonstrated. Using the triple complex formation to facilitate DEP separation, a method is developed to detect B/F binding by a direct observation of the separation pattern on the microelectrode system. Et is applied for: (1) quantitative detection of alpha-fetoprotein, the diagnostic marker of liver cancer, through antigen-antibody reaction; and (2) the detection of DNA sequence through hybridization. The methods developed here are compatible with with micro fabrication, and suitable for affinity assays in micro-total analysis systems.