Simultaneous multiple analyte detection using fluorescent peptides and capillary isoelectric focusing.

Analyte-specific detection based on the isoelectric point of the detection moiety is a new concept that is under investigation at Vysis. We have developed methods for the synthesis of of fluorescent synthetic peptides that can be conjugated to bioanalytes such as nucleic acids and antibodies, processed in a hybridization or binding assay, and then chemically released prior to detection by capillary isoelectric focusing (cIEF)-laser-induced fluorescence (LIF) detection. A two-step cIEF method in coated capillaries using salt mobilization has been used that produces high peak efficiencies and good assay reproducibility. The concentration by focusing aspect of cIEF, which allows for the entire capillary to be filled with sample, enables detection limits in the pM as opposed to sub-nM level for conventional capillary electrophoresis (CE)-LIF. The simultaneous multiple detection of eleven different focusing entities has been achieved.