K. Adachi et al., Single-fluorophore imaging with an unmodified epifluorescence microscope and conventional video camera, J MICROSC O, 195, 1999, pp. 125-132
Single fluorophores in aqueous solution were imaged in real time with a con
ventional silicon-intensified target video camera connected to an unmodifie
d commercial microscope (IX70, Olympus) with epifluorescence excitation wit
h a high-pressure mercury lamp. Neither a powerful laser nor an extremely s
ensitive video camera was required, Three experimental systems were used to
demonstrate quantitatively that individual, moving or stationary Cy3 fluor
ophores could be imaged with the microscope: Cy3-gelsolin attached to an ac
tin filament sliding over heavy meromyosin, sliding actin filaments sparsel
y labelled with Cy3, and heavy meromyosin labelled with one or two Cy3 fluo
rophores. The results should encourage many laboratories to attempt 'single
-molecule physiology' in which the functions and mechanisms of molecular ma
chines are studied at the single-molecule level in an environment where the
biological machines are fully active.