Technology Review: Introduction

The inception of the human genome project created a tremendous need for fast, reliable and low cost automated DNA sequencing. LI-COR was one of the pioneers of automated DNA sequencing based on visible fluorescence detection. However, the intrinsic fluorescence of biomolecules posed a problem when visible light absorbing fluorophores were used. Near-infrared (NIR) fluorescence (approx. 670-1000 nm) detection, by contrast, avoided the background fluorescence interference of natural biomolecules as well as polyaromatic hydrocarbons. The NIR region was also compatible with solid-state optical components such as diode lasers and silicon detectors.

Instead of commercializing a DNA sequencer based on visible fluorescence, LI-COR developed a new NIR fluorescence detection system and high quantum yield NIR dyes (now called IRDye® infrared dyes). This new detection system, first commercialized in 1993, had inherent advantages due to NIR detection, such as wide dynamic range and very high sensitivity. The resulting DNA sequencer, and its pioneering use of NIR technology, set new standards for readlength and accuracy.

Today, LI-COR uses infrared detection systems in its Model 4300 DNA Analysis System, Odyssey® Infrared Imaging System and Aerius® Automated Infrared Imaging System. Each system uses two channel detection with matched IRDye® Infrared Dyes for a wide variety of applications including Microsatellite analysis, Tilling®, Westerns, In-Cell Westerns, In Vivo imaging, and tissue analysis.

IR Advantages include...

• Wider dynamic range and minimal background as a result of reduced scattering compared to visible fluorescence detection (due to the inverse 4th power dependence on the wavelength).
• Very high sensitivity resulting from low infrared background, and the high extinction coefficients of IRDye® infrared dyes.
• High sensitivity lowers reagent usage and enables smaller detection geometries.
• Low cost diode laser excitation sources and cooled silicon avalanche photodiodes (APD) for detection reduce power consumption and make IR fluorescence systems more economical and compact.
• Solid-state IR diode lasers and APDs have unparalleled reliability and the lowest maintenance costs.
• IR fluorescence detection can replace radioactivity or indirect detection methods (such as enzyme-based chemiluminescent or colorimetric detection) for applications requiring blotting on membranes due to low autofluorescence from membranes and other biomolecules providing greater sensitivity than can be achieved using shorter-wavelength visible fluorophores.

One or more LI-COR patents may cover IRDye® infrared dyes, including the following: US06027709; and several other pending patents.