Label-free characterization of cell adhesion using reflectometric interference spectroscopy (RIfS)

Abstract

Reflectometric interference spectroscopy (RIfS) is a label-free, time-resolved technique for detecting interactions of molecules immobilized on a surface with ligands in solution. Here we show that RIfS also permits the detection of the adhesion of tissue culture cells to a functionalized surface in a flow system. Interactions of T cells with other leukocytes or epithelial cells of blood vessels are crucial steps in the regulating immune response and inflammatory reactions. Jurkat T cell leukemia cells rapidly attached to a transducer functionalized with a monoclonal antibody directed against the T cell receptor (TCR)/CD3 complex, followed by activation-dependent cell spreading. RIfS curves were obtained for the Jurkat derivative JCaM 1.6 (which lacks the key signaling protein Lck), cells preincubated with cytochalasin D (an inhibitor of actin polymerization), and for surfaces functionalized with an antibody directed against the coreceptor CD28. These curves differed with respect to the maximum signal and the initial slope of the increase in optical thickness. The testing of chemical inhibitors, cell surface molecules and gene products relevant to a key event in T cell immunity illustrates the potential of label-free techniques for the analysis of activation-dependent cell-surface contacts.

Diet-Derived Polyphenol Metabolite Enterolactone Is a Tissue-Specific Estrogen Receptor Activator

“…4A). RIfS is a label-free, time-resolved sensing method to measure biomolecular interactions, which has been applied to various applications (31)(32)(33). The RIfS set-up is described in detail in supplemental Fig.…”

Direct Optical Detection in Bioanalytics

“…To qualify the surface modification, affinity constants of LNA (locked nucleic acid) and DNA duplex formation were determined [211] and endocrine receptors were examined [212];cell morphology also can be examined by RIfS [213]. Binding of ubiquitin to short peptide segments of hydrolase was studied in comparison to fluorescence correlation spectroscopy, isothermal calorimetry, and NMR [214].…”

Spectral Self-Interference Fluorescence Microscopy to Study Conformation of Biomolecules with Nanometer Accuracy

“…WLRS offers great precision for determination of the thickness of a transparent spacer layer on a reflecting surface [63][64][65][66][67][68][69]. A broadband light source is focused onto the surface and reflected light is collected.…”