Development of a micro total analysis system incorporating chemiluminescence detection and application to detection of cancer markers.

We developed a micro total analysis system (mu-TAS) incorporating chemiluminescence detection, in which the chemiluminescence reaction of isoluminol isothiocyanato (ILITC) (as a chemiluminescence reagent for labeling)-microperoxidase (as a catalyst)-hydrogen peroxide (as an oxidant) was adopted. The analysis system performed the following three processes on a microchip: immune reaction for high selectivity, electrophoresis for formation and transportation of the sample plug, and chemiluminescence detection for high sensitivity. The three processes were compactly integrated onto the microchip to give the mu-TAS. The microchip contained two microchannels that crossed at an intersection, while the ends of the microchannels accessed four reservoirs. As the first process, the immune reaction was performed using an antibody-immobilized glass bead. The glass bead was placed in one of the reservoirs along with antigen (analyte) and a known amount of ILITC-labeled antigen to set up a competitive immune reaction. For electrophoresis, as the second process, the reactant after the immune reaction was fed electrophoretically into the intersection resulting in a sample plug. The sample plug was then moved into another reservoir containing hydrogen peroxide solution. At this point, chemiluminescence detection was performed as the third process: the labeled antigen mixed with the hydrogen peroxide and the catalyst included in the migration buffer to produce chemiluminescence. Chemiluminescence was detected by a photomultiplier tube located under the reservoir. The mu-TAS described here was capable of determining, with high selectivity and sensitivity, human serum albumin or immunosuppressive acidic protein as a cancer marker in human serum.