Abstract
BackgroundA critical challenge in cell biology is quantifying the interactions of cells with their extracellular matrix (ECM) environment and the active remodeling by cells of their ECM. Fluorescence microscopy is a commonly employed technique for examining cell-matrix interactions. A label-free imaging method would provide an alternative that would eliminate the requirement of transfected cells and modified biological molecules, and if collected nondestructively, would allow long term observation and analysis of live cells.ResultsUsing surface plasmon resonance imaging (SPRI), the deposition of protein by vascular smooth muscle cells (vSMC) cultured on fibronectin was quantified as a function of cell density and distance from the cell periphery. We observed that as much as 120 ng/cm2 of protein was deposited by cells in 24 h.ConclusionSPRI is a real-time, low-light-level, label-free imaging technique that allows the simultaneous observation and quantification of protein layers and cellular features. This technique is compatible with live cells such that it is possible to monitor cellular modifications to the extracellular matrix in real-time.
Highlights
A critical challenge in cell biology is quantifying the interactions of cells with their extracellular matrix (ECM) environment and the active remodeling by cells of their ECM
These results demonstrate the lateral resolution of the surface plasmon resonance imaging (SPRI) system to be at least 2 Pm based on the ability to distinguish 2 Pm features spaced 2 Pm apart
Our results show that Surface plasmon resonance (SPR) images, obtained without the addition of fluorescent or other labels, can provide information on cell-substrate contacts, ECM deposition and substrate protein patterns
Summary
A critical challenge in cell biology is quantifying the interactions of cells with their extracellular matrix (ECM) environment and the active remodeling by cells of their ECM. A label-free imaging method would provide an alternative that would eliminate the requirement of transfected cells and modified biological molecules, and if collected nondestructively, would allow long term observation and analysis of live cells. We show here that as an alternative, SPRI can be a sensitive, label-free, and low-light optical method that eliminates the requirement for modified biological molecules and transfected cells, and allows for highly sensitive real-time observation of protein deposition and live cell engagement with the ECM. Because the refractive index is proportional to the amount of adsorbate at the surface [7], SPR has been used as a quantitative, sensitive, and label-free technique for measuring the binding kinetics of proteins [8], DNA [9,10], and small molecules [11,12], to surface immobilized capture agents. SPR imaging has not previously been considered a useful technique for imaging cell features, largely because of previous assumptions that poor spatial resolution would prevent useful imaging
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