The development of novel techniques to accurately deliver, manipulate, and detect single molecules is one of the most interesting but challenging subjects in modern biotechnology and molecular biology. Most of the recent studies are focused on electrochemical methods using glass micro/nano pipettes that are mechanically pulled and an electrical field used to move molecules through microchannels. However, “single-molecule delivery” has not yet been achieved.

We report on our recent studies of single molecule delivery using a sub-attoliter reservoir formed at the end of what is essentially an NSOM optical fiber, mounted on a combined AFM/confocal fluorescence microscope. The reservoir is formed by chemical etching and focused ion beam (FIB) milling and is then coated with Al to confine the light for fluorescence detection in an NSOM geometry, and for the expulsion of the solution containing the molecule of interest. The volume of the reservoir can be in the range of attoliters by control of the etching and/or milling conditions. Initial experimental results using fluorescein dye indicates that when the solution concentration is low enough, single molecule delivery is achievable. Further study on cell labeling is discussed.

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