Wavelength-Dependent Differential Interference Contrast Microscopy: Multiplexing Detection Using Nonfluorescent Nanoparticles

The wavelength dependence of plasmonic nanoparticles’ contrasts in differential interference contrast (DIC) microscopy has been exploited previously for unambiguous identification and dynamic tracking of these nanoprobes in complex environments (Anal. Chem. 2009, 81, 9203−9208). In the present study, the suitability of multiplexing detection in DIC microscopy was investigated systematically with 19 kinds of nanoparticles of different materials and/or sizes. A unique DIC contrast spectrum was found for each kind of nanoparticle. Multiplexing detection was accomplished by measuring DIC contrasts at a minimum of two specific illumination wavelengths. The main advantages of DIC microscopy for multiplexing detection over other nonfluorescence techniques, such as dark field microscopy and surface-enhanced Raman scattering, were demonstrated by differentiating four kinds of nanoparticles on the cell membrane while providing high-contrast images of both the nanoprobes and cell features.