Applications of High Resolution Surface Plasmon Resonance Imaging to Adherent Cells: Single Mammalian Cells to Bacterial Biofilms

Abstract

High resolution surface plasmon resonance imaging (SPRI) allows label-free imaging of subcellular features when performed using a high numerical aperture objective lens with a digital light projector to precisely position incident angle excitation. The SPRI signal is a result of the mass of material within the evanescent field, and can be interpreted as a density or a refractive index of the cellular material. The quantitative nature of SPR images and the direct relationship to refractive index changes at the surface sensor allow for visualization new insights into mechanisms of cell biology at an interface. When applied to mammalian cells, such as rat aortic smooth muscle cells, cellular components near the sensor surface such as the cell membrane, focal adhesions, and cell nucleus are visualized in the SPRI image. Focal adhesion sizes measured by SPRI are similar with those highlighted with fluorescent antibody stained vinculin. In addition, a positive correlation between focal adhesion size and protein density is observed by SPR imaging. When SPRI is applied to pathogenic biofilms of Streptococcus mutans, distinct components of the bacterial biofilm at the surface including individual bacteria, bacterial microcolonies, and extracellular polymeric substance (EPS) are observed. SPRI shows that the refractive index of bacteria in a biofilm increases over time compared to that of bacteria not in a biofilm, which remains constant. SPRI also indicates that the EPS material generated in the biofilm at early time points is thicker near the bacterial microcolony periphery. This suggests that the EPS matrix is generated at the colony edge and that SPRI can be used to monitor the dynamics of EPS production in biofilms.