Vibrational spectra of individual millimeter-size membrane patches using miniature infrared waveguides.
We have used miniature planar IR waveguides, consisting of Ge strips 30-50 microm thick and 2 mm wide, as evanescent-wave sensors to detect the mid-(IR) evanescent-wave absorbance spectra of small areas of biomolecular monolayers and multilayers. Examples include picomolar quantities of an integral transmembrane protein (bacteriorhodopsin) and lipid (dimyristoyl phosphatidylcholine). IR bands due to the protein and lipid components of the plasma membrane of individual 1.5-mm-diameter devitellinized Xenopus laevis oocytes, submerged in buffer and sticking to the waveguide surface, were also detected. A significant improvement in sensitivity was observed, as compared to previous sizes and geometries of evanescent-wave sensors (e.g., commercially available internal reflection elements or tapered optical fibers). These measurements suggest the feasibility of using such miniature supported planar IR waveguides to observe structural changes in transmembrane proteins functioning in vivo in single cells.
Plunkett
S E
SE
University of Virginia Health Sciences Center, Department of Biochemistry, Charlottesville 22908, USA.
Jonas
R E
RE
Braiman
M S
MS
D000818:Animals; D001436:Bacteriorhodopsins / Q000737:chemistry; D055592:Biophysical Phenomena; D001703:Biophysics; D002462:Cell Membrane / Q000737:chemistry*; D004134:Dimyristoylphosphatidylcholine / Q000737:chemistry; D005260:Female; D066298:In Vitro Techniques; D008563:Membrane Lipids / Q000737:chemistry; D008565:Membrane Proteins / Q000737:chemistry; D009865:Oocytes / Q000737:chemistry; D017550:Spectroscopy, Fourier Transform Infrared / Q000379:methods*; D014982:Xenopus laevis
D008563:Membrane Lipids; D008565:Membrane Proteins; D001436:Bacteriorhodopsins; D004134:Dimyristoylphosphatidylcholine