Immobilization of antibodies in micropatterns for cell detection by optical diffraction

Optical diffraction at biochemically microstructured surfaces has been investigated for the label-free in situ detection of cells. The new sensor concept is based on regular arrays of covalently coupled antibodies, which selectively bind cells from solution. Due to the adsorption process, changes are imposed on the intensity distribution of the diffracted light, which can serve to quantify the amount of adsorbed cells. For the formation of such microstructures, different classical film preparation techniques were transferred to a mesoscopic scale by the use of microcontact printing (μCP). Alternatively, receptors were functionalized with thiol groups prior to the immobilization process and directly printed onto the gold surface. Compared to imprinting of non-functionalized proteins on gold, a better replication of the micropatterns could be obtained. Additionally, a significantly lower amount of defects was observed than for the classical coupling techniques. Using such microstructures, first experiments on the detection of Escherichia coli bacteria were performed. Diffraction patterns have been observed for concentrations equal or higher than 106 cells/ml. In time dependent experiments, diffraction spots occurred after 30 – 90 min or 10 – 20 min, depending on whether non-specific cell adsorption or specific binding to anti-E. coli IgG was studied. A first quantitative analysis of the diffraction patterns shows that the total amount of diffracted light increases with increasing incubation time.