Fabrication of Protein Microarrays for Immunoassay Using the Electrospray Deposition (ESD) Method

In recent years new methods for active protein film fabrication have been rapidly developing. One of these methods is the electrospray deposition (ESD) method. This method allows the spontaneous deposition of many identical dots and has a remarkable spatial resolution as well as the ability to overcome limitations like low rates of deposition, low efficiency of substance transfer, and cross-contamination. In this study, an antibody-based protein array for high-throughput immunoassay was fabricated with the ESD method using a quartz mask with holes made by an abrasive jet technique. An antibody solution was electrosprayed onto an ITO glass and then deposited antibodies were cross-linked with a vapor of glutaraldehyde. The diameter of the spots was approximately 150 μm. The arrays were then incubated with corresponding target antigenic molecules and washed. The captured antigens were collectively detected by fluorescence and chemiluminescence. These signals were quantitatively visualized with a high-resolution charge-coupled device (CCD) detector. Sensitive (~1 ng/ml) and simultaneous detection of various antigens could be performed by enzyme linked immunosorbent assay or fluorescence immunoassay using this system.