Braun98

Braun98 - VOLUME 81, NUMBER 23 P H Y S I C A L R E V I E W...

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Unformatted text preview: VOLUME 81, NUMBER 23 P H Y S I C A L R E V I E W L E T T E R S 7 D ECEMBER 1998 Fluorescence Interferometry of Neuronal Cell Adhesion on Microstructured Silicon Dieter Braun and Peter Fromherz* Department Membrane and Neurophysics, Max-Planck-Institute for Biochemistry, D-82152 Martinsried/Mnchen, Germany (Received 8 July 1998) We measured the separation of cell membranes from a surface of silica with nanometer precision taking advantage of the fluorescence of an organic dye in the standing modes of light above silicon. For neural cells from rat brain we found about 105 nm on a surface coated with laminin and about 60 nm with fibronectin. No plaques of close adhesion were seen within a lateral resolution of 400 nm. The wide homogeneous cleft raises the question about the nature of the attractive and repulsive forces in cell adhesion. [S0031-9007(98)07867-3] PACS numbers: 87.22.Bt, 07.60.Ly, 33.50.Dq, 78.66.w Migration and outgrowth of living cells are investi- gated generally on solid substrates coated with proteins of the extracellular matrix [13]. Little is known about the microscopic structure of the cell adhesion which is the basis for any description on a molecular level. We mea- sured the distance of the membrane of neuronal cells from a silica surface coated with laminin and fibronectin, taking advantage of the fluorescence of a membrane-bound dye in the standing light waves above a microstructured sili- con chip. The method is insensitive to ill known optical parameters of the cell, yielding a systematic error of about 1 nm. We found a surprisingly wide extracellular cleft of 105 and 60 nm on laminin and fibronectin, respectively, which was surprisingly homogeneous within 5 nm with- out visible plaques of close adhesion. Standing waves of the electromagnetic field form in front of a reflecting interface. The fluorescence intensity of a dye molecule depends on the distance from the interface due to a modulation of excitation and emission [47]. We consider a cell grown across microscopic steps of silicon dioxide (height D d ox 20 nm, width 2.5 m m ) on silicon (Fig. 1) with a distance d cleft between cell membrane and chip. The membrane is placed by the steps at different positions in the standing waves. If it is stained with a dye, we expect a pattern of fluorescence intensity in register with the steps. The intensity as a function of the thickness of oxide d ox depends on the distance d cleft . Vice versa d cleft can be obtained by fitting experimental data by a multireflection optical theory [8,9]. This method of fluorescence interference contrast (FLIC) microscopy was tested in model systems such as Langmuir-Blodgett films [8] and erythrocyte membranes [9]. It relies on homogeneous staining, illumination, and sensitivity of detection....
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Braun98 - VOLUME 81, NUMBER 23 P H Y S I C A L R E V I E W...

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