Abstract

Single dye molecules may be detected as they pass through a 0.4 pL sample volume by using repetitively pulsed laser excitation and time-gated single photon counting to detect the weak single-molecule fluorescence and simultaneously discriminate promptly scattered Raman photons arising from the >10¹³ solvent molecules.¹ Previous experiments have employed a microchannel plate photomultiplier (MCP), which achieves good time response but mediocre quantum efficiency. For comparison, we have characterized the performance of a single photon avalanche diode (EG&G SPCM-200 series) custom modified to allow the avalanche pulses to be directly coupled to fast timing modules. For a reverse bias of 20 V above breakdown a considerably better overall photon detection efficiency of ~50% is achieved, with a dark noise of 60 s⁻¹ when the sample is thermoelectrically cooled to about −45°C. Since the sample volume in the single molecule detection experiment is very small, the collected light may be tightly focused onto the detector. When care is taken to focus to a 10 μm spot correctly positioned on the diode, the time response for time-gated photon counting is comparable with or better than that obtained by the MCP. Although the prompt full-width-at-half-maximum (FWHM) is longer (168 ps), the absence of optical reflections results in a prompt with a shorter tail [FW(1/100)M = 790 ps], which allows the time gate to begin at earlier times.

© 1992 Optical Society of America