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Multiphoton plasmon-resonance microscopy

Dvir Yelin, Dan Oron, Stephan Thiberge, Elisha Moses, and Yaron Silberberg

Open Access Open Access

Abstract

A novel method for detection of noble-metal nanoparticles by their nonlinear optical properties is presented and applied for specific labeling of cellular organelles. When illuminated by laser light in resonance with their plasmon frequency these nanoparticles generate an enhanced multiphoton signal. This enhanced signal is measured to obtain a depth-resolved image in a laser scanning microscope setup. Plasmon-resonance images of both live and fixed cells, showing specific labeling of cellular organelles and membranes, either by two-photon autofluorescence or by third-harmonic generation, are presented.

©2003 Optical Society of America

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Figures (4)
Fig. 1.
Fig. 1. Plasmon-resonance TPAF images of CHO cells incubated with (a) 10nm cationic gold. (b) 40nm Concanavalin A gold. (c) two cells which were submitted to the same treatments performed in the positive experiments (a and b), but no gold was added to the growing medium. The strong TPAF spots in a and b are attributed to plasmon-resonance with the laser frequency where nanoparticles are aggregated.

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Fig. 2.
Fig. 2. Plasmon-resonance TPAF image of a live CHO cell incubated with 10nm cationic gold, superimposed on a simple transmission image of the cell. The transmission image is blue, while the TPAF signal goes from red (weak) to yellow (strong).

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Fig. 3.
Fig. 3. (a) THG image of a fixed NIH3T3 cell. While the nucleus appears dark, numerous bright spots are observed in the cell volume. (b) THG image of a NIH3T3 cell, in which the nucleus membrane was labelled by 10nm gold nanoparticles followed by silver enhancement.

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Fig. 4.
Fig. 4. Four THG sections of a NIH3T3 epithelial cell whose membrane was labelled by 10nm gold nanoparticles, followed by silver enhancement. Bright areas denote strong third-harmonic signal. The top of the cell is shown on the top left corner, while the bottom of the cell is shown on the bottom right corner. The spacing between sections is approximately 3µm. The average illumination power is of the order of 1mW. The cell shape, in the form of a triangular pyramid is clearly evident. The inside of the cell, where there are no nanoparticles, is completely dark.

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