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March 01, 2009 (Vol. 29, No. 5)

Drug Absorption on Membrane Arrays

A Novel Method Using SPR Imaging

  • SPR Imaging Technology

    The SPRimager®II from GWC Technologies measures binding events that occur on the surface of a gold-coated chip. Binding of an analyte to probe molecules immobilized on a chip results in an increase in reflectivity (D%R). A CCD camera captures this change in reflectivity by imaging the whole array in real time. 

    In a typical experiment, a reference image of the array is collected prior to exposure to analyte. Then, images are collected as the array is exposed to a flow of analyte solution. For each time point, the reference image is subtracted and D%R for each element on the array is plotted as a function of time. An increase in reflectivity indicates binding has occurred. A decrease in reflectivity indicates dissociation.

  • Fabrication of Arrays

    Click Image To Enlarge +
    Figure 1A and 1B. Membrane array evaluation

    Drug-lipid interactions have been measured in a label-free manner using synthetic lipids in the form of liposomes. In this assay, natural cell membranes from Spodoptera frigiperda Sf9 insect cells (Invitrogen) were immobilized on gold-coated SPRi chips. The core of this approach is the incorporation of biotinylated PEG-phospholipids into the membrane (using double sonication) to generate biotin-tagged, small membrane vesicles. 

    Membrane fractions of Sf9 insect cells were sonicated with polymeric micelles composed of biotin-PEG-DSPE (Avanti Polar Lipids). After washing and centrifugation, the supernatant containing the biotinylated cell membranes (biMEM) was diluted and spotted on streptavidin-coated SPRi sensors where they were immobilized via the biotin-containing anchors.  

    An image of a typical biMEM array on a SpotReady16™ chip before exposure to analyte is shown in Figure 1A. biMEM capture on the chip surface is specific, and probe density of tethered membranes increases as the concentration of spotted biMEM increases (Figure 1B).