High-performance immunoassays based on through-stencil patterned antibodies and capillary systems

We present a simple method to pattern capture antibodies (cAbs) on poly(dimethylsiloxane) (PDMS), with high accuracy and in a manner compatible with mass fabrication for use with capillary systems (CSs), using stencils microfabricated in Si. Capture antibodies are patterned as 60 - 270 mu m wide and 2 mm long lines on PDMS and used with CSs that have been optimized for convenient handling, pipetting of solutions, pumping of liquids, such as human blood serum, and visualization of signals for fluorescence immunoassays. With the use of this method, C-reactive protein (CRP) is detected with a sensitivity of 0.9 ng mL(-1) (7.8 pM) in 1 mu L of CRP-spiked human serum, within 11 min and using only four pipetting steps and a total volume of sample and reagents of 1.35 mu L. This exemplifies the high performances that can be achieved using this approach and an otherwise conventional surface sandwich fluorescence immunoassay. This method is simple and flexible and should therefore be applicable to a large number of demanding immunoassays.