Advances in Detector Systems for Imaging Single Molecule Fluorescence

✍ Scribed by Mark Christenson

• Publisher: John Wiley and Sons
• Year: 2000
• Tongue: English
• Weight: 97 KB
• Volume: 1
• Category: Article
• ISSN: 1438-5163
• DOI: 10.1002/1438-5171(200006)1:2<177::aid-simo177>3.0.co;2-4

✦ Synopsis

In order to image single molecule fluorescence (SMF), an optimized optical microscope station is required as well as a high sensitivity detection system. This talk describes the current trends in detectors for SMF using back illuminated CCD technology as well as image intensified technology. A comparison of the two different detection methodologies will be presented as well as some recent advances in very high speed, low light detection methods.

The optimal detector for single molecule fluorescence will have 100% photo-conversion efficiency and zero system noise. While this detector does not exist, there are devices that approach these specifications under restricted operating conditions. Back illuminated CCDs have the ability to convert approximately 80-90% of the visible photons (500-650 nm) hitting the surface of the device (Figure ). This generates the high signal required for sensitivity. In order to prevent a dark current background signal from being generated within the CCD itself, an efficient cooling mechanism is required to lower the thermal energy of the CCD. If this is done properly, the dark current is essentially zero for the duration of the exposure and readout.

Now the accumulated signal on the CCD can be digitized with very low noise (3-4 electrons rms). There is, however,