Combined laser tweezers and dielectric field cage for the analysis of receptor-ligand interactions on single cells.
A new technique based on the combination of optical and chip-based dielectrophoretical trapping was developed and employed to manipulate cells and beads with micrometer precision. The beads were trapped with optical tweezers (OT) and brought into contact for defined times with cells held in the dielectrophoretic field cage (DFC). The well-defined ligand-receptor system biotin-streptavidin was used to study the multiple interaction between biotinylated live cells and streptavidin-coated beads. The biotin density on the cell surface was varied down to a few single bonds (3 +/- 2 bonds/microm2) to control the valency of the binding. The quantitative relationship between the contact area, ligand density and its diffusion rate in the outer membrane of the cell could be demonstrated. The increase of the strength of the cell-bead adhesion was strictly dependent on the increase of individual bond numbers in the contact area. This is in part due to accumulation of ligands (D approxiamtely (0.5 +/- 0.1) 10(-8) cm2/s) in the contact area as seen by confocal laser scanning microscopy. Individual receptor-ligand rupture forces were evaluated and are compatible with values obtained by biomembrane force probe techniques. To summarize, the combination leads to a new powerful microsystem for cell handling and pN-force measurements on the single-cell level.
Reichle
C
C
Department of Biology/Biophysics, Humboldt University, Berlin, Germany.
Sparbier
K
K
Müller
T
T
Schnelle
T
T
Walden
P
P
Fuhr
G
G
D000818:Animals; D001710:Biotin / Q000378:metabolism; D019921:Biotinylation; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D004574:Electromagnetic Fields; D004867:Equipment Design; D007834:Lasers; D008024:Ligands; D046911:Macromolecular Substances; D051379:Mice; D008810:Mice, Inbred C57BL; D008846:Micromanipulation / Q000295:instrumentation*; D008854:Microscopy, Electron; D008856:Microscopy, Fluorescence; D008863:Microspheres; D011485:Protein Binding; D011956:Receptors, Cell Surface / Q000378:metabolism*; D019809:Streptavidin / Q000378:metabolism; D013945:Thymoma / Q000473:pathology; D013953:Thymus Neoplasms / Q000473:pathology; D014407:Tumor Cells, Cultured / Q000378:metabolism / Q000648:ultrastructure
D008024:Ligands; D046911:Macromolecular Substances; D011956:Receptors, Cell Surface; D001710:Biotin; D019809:Streptavidin