Combination of AFM with an objective-type total internal reflection fluorescence microscope (TIRFM) for nanomanipulation of single cells.

A new instrument was constructed by combining an objective-type total internal reflection fluorescence microscope with an atomic force microscope (AFM). Our purpose of constructing such an instrument is to detect and confirm the result of cellular level manipulations made with the AFM part through the detection system of the highly sensitive fluorescence microscope part. In this combination, manipulations are now possible from the nanometer to the micrometer scales and the fluorescence detection system is sensitive enough even for localizing single molecules. In this paper, we applied the system as a precise intracellular injector (nanoplanter). Fluorescent beads were first chemically immobilized onto a ZnO whisker that was glued to an AFM tip and were injected into a living BALB/3T3 cell together with the whisker. It was demonstrated that the system could clearly show the result of injection, that is, the presence of a small number of fluorescent beads in the cell.