The second law of thermodynamics sets a fundamental limit to the efficiency with which a large, linear array of capillaries can be detected. Briefly, a magnifying optical system, such as a microscope, can image light efficiently. However, a demagnifying system suffers from an inherent loss of light that is proportional to the square of the demagnification factor. Demagnification occurs when the size of the linear array exceeds the size of the detector array. For example, a 2-cm capillary array would be demagnified by a factor of two when imaged onto a 1-cm CCD chip. This demagnification becomes important for arrays that contain several hundred capillaries.
        We developed an optical system that packs a very large number of capillaries into a small space, which allows for efficient detection of fluorescence. In this two-dimensional direct reading spectrograph, the capillaries (marked 2 in the figure) are held in a two-dimensional bundle, much as pencils are held in a box. A set of thin metal shims has an array of holes matched to the outer dimension of the capillaries (3). The capillaries are threaded through the shims to align them. The shims themselves are held in place by two precision milled posts (1). Another shim (4), called a barrier member, is located a few hundred micrometers down stream from the capillary array. Buffer is pumped from the inlet (6) above the barrier member and exits (7) beneath the member. As analyte exits the capillary tips, it is entrained in the flowing buffer and forced to pass through the corresponding hole in the barrier. An elliptically-shaped laser beam (5) skims above the barrier member to excite fluorescence from analyte. A window (9) is provided beneath the array to allow imaging of fluorescence through the holes in the member.

The barrier member is shown below. We had a number of these plates machined from nickel, and they serve to both align the capillaries and to provide flow path for analyte.

        We developed a two-dimensional direct reading spectrograph for this instrument to monitor the emission spectrum from each sample stream. A camera-lens collimates fluorescence from each sample stream; the camera images fluorescence through the window (9) in the bottom of the cuvette. A prism disperses the spectrum, which is re-imaged onto the face of a CCD camera. We used a prism, rather than a grating, to eliminate inter-capillary cross-talk by eliminating higher-order spectra, which would otherwise overlap with the spectrum of adjacent capillaries.

        The image from each capillary appears as a spectrum. The 96-spectra are tightly packed, creating a rather pretty view. Note that by offsetting the holes in the alignment plates, we provide additional space for the spectra, taking advantage of the available real-estate on the CCD camera.

        We record the fluorescence intensity from each pixel, and use that information to reconstruct the emission spectrum of analyte migrating from each capillary. This data is then used to generate a four-color DNA electropherogram.
        The instrument has been described in the paper "Two-Dimensional Direct-Reading Fluorescence Spectrograph for DNA Sequencing by Capillary Array Electrophoresis" J. Zhang, M. Yang, X. Puyang, Y. Fang, L.M. Cook, N.J. Dovichi, Analytical Chemistry 73, 1234-1239 (2001).