University of Regensburg Trenner Faculty of Chemistry Trenner Analytical Chemistry
Single molecule analysis in femtoliter arrays

Single enzyme molecule detection in femtoliter array

Glass fiber bundles with a diameter of 2 mm consisted of 50,000 fiber cores (white) embedded in a common cladding (gray). Due to the different material composition of core and cladding, the fibers could be etched specifically with HCl to generate a homogeneous array of femtoliter containers on the distal surface of the fiber bundle. Each container in the array was optically “wired” to its adjacent fiber. As the fiber core material had a higher refractive index than the cladding material light was totally internally reflected in the fibers that served as waveguides for excitation and emission light.

The fiber bundle was mounted on an upright epifluorescence microscope and the reaction monitored through the proximal side (p) of the fiber bundle after sealing the femtoliter-containers with a silicone gasket (gasket not shown). The individual β-galactosidase molecules in the femtoliter-containers converted the non-fluorescent substrate resorufin-β-galactopyranoside to fluorescent resorufin (yellow containers). The substrate turnover could be temporarily interrupted by the slow-binding inhibitor D-galactal.

(FEBS Journal 2007)

© 2014, Thomas Hirsch, Gisela Emmert.