Protein microarrays imaging using oblique-incidence reflectivity difference method

In this study Oblique-Incidence Reflectivity Difference (OIRD) technique, a recent developed label-free detection, is applied to image biomolecular microarrays. Compared to the currently widely used fluorescence-dependent optical microscopy, OIRD technique not only images the morphology of protein bio-arrays in the absence of extrinsic labeling molecules but also monitors the changes in the optical properties of biochips in high-throughput fashion. Additionally, such a technique complements other label-free detections including Surface Plasmon Resonance (SPR) and Mass Spectrometry (MS) by offering the opportunity to detect biochemical activities without the special requirements on the substrate or the specific matrix medium. It is shown in this article that the surface topography can be reflected by OIRD method. Besides, the differences among printing concentrations and various proteins are able to be identified as well. Incidentally, the OIRD images appear to be useful aids to distinguish the variations resulting from antibody-antigen capture. Both the imaginary and real parts of OIRD images we presented provide more information than the single imaginary one especially. On the basis of the scattering mechanism, the absorptive properties of protein molecules are analyzed through the real part of OIRD signal. Accordingly, OIRD technique shows its unique potential in detection of biochemical processes.